PI: LOCHHEAD, MICHAEL J Title: Point-of-Care HIV Antigen/Antibody Diagnostic Device

Received: 05/07/2013 FOA: PA13-088 Council: 10/2013

Competition ID: ADOBE-FORMS-B2 FOA Title: PHS 2013-02 Omnibus Solicitation of the NIH, CDC, FDA and ACF for

Small Business Innovation Research Grant Applications (Parent SBIR [R43/R44])

2 R44 AI093289-02A1 Dual: Accession Number: 3586675

IPF: 10026307 Organization: MBIO DIAGNOSTICS, INC.

Former Number: Department:

IRG/SRG: ZRG1 AARR-E (81)B AIDS: N Expedited: N

Subtotal Direct Costs Animals: N New Investigator: N

(excludes consortium F&A) Humans: Y Early Stage Investigator: N

Year 2: Clinical Trial: N

Year 3: Current HS Code: 20

HESC: N

Senior/Key Personnel: Organization: Role Category:

Michael Lochhead MBio Diagnostics, Inc. PD/PI 

Daniel Nieuwlandt MBio Diagnostics, Inc. Other (Specify)-Senior Scientistg

Constance Benson University of California, San Diego Co-Investigator

Susan Little University of California, San Diego Co-Investigator

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State Application Identifier

Applicant Identifier

1. * TYPE OF SUBMISSION 4. a. Federal Identifier

5. APPLICANT INFORMATION * Organizational DUNS:

* Legal Name:

Department: Division:

* Street1:

Street2:

* City:

* State:

* Country: * ZIP / Postal Code:

Person to be contacted on matters involving this application

* First Name: Middle Name:

* Last Name: Suffix:

* Phone Number: Fax Number:

Email:

6. * EMPLOYER IDENTIFICATION (EIN) or (TIN):

7. * TYPE OF APPLICANT:

Other (Specify):

Small Business Organization Type Women Owned Socially and Economically Disadvantaged

If Revision, mark appropriate box(es).

9. * NAME OF FEDERAL AGENCY:

A. Increase Award B. Decrease Award C. Increase Duration D. Decrease Duration

E. Other (specify):

10. CATALOG OF FEDERAL DOMESTIC ASSISTANCE NUMBER:

* Is this application being submitted to other agencies?

TITLE:

2. DATE SUBMITTED

3. DATE RECEIVED BY STATE APPLICATION FOR FEDERAL ASSISTANCE

SF 424 (R&R) 

County / Parish:

Province:

Prefix:

What other Agencies?

Pre-application Application Changed/Corrected Application

MBio Diagnostics, Inc.

5603 Arapahoe Avenue,Suite 1

Boulder `

80303-1377

Cynthia

Moore

11. * DESCRIPTIVE TITLE OF APPLICANT'S PROJECT:

Point-of-Care HIV Antigen/Antibody Diagnostic Device

Yes No

USA: UNITED STATES

CO: Colorado

R: Small Business

National Institutes of Health

New Resubmission

Renewal Continuation Revision

8. * TYPE OF APPLICATION:

OMB Number: 4040-0001 

Expiration Date: 06/30/2011 

b. Agency Routing Identifier

12. PROPOSED PROJECT:

* Start Date * Ending Date

10/01/2013 09/30/2015

* 13. CONGRESSIONAL DISTRICT OF APPLICANT

CO-002

14. PROJECT DIRECTOR/PRINCIPAL INVESTIGATOR CONTACT INFORMATION

* First Name: Middle Name:

* Last Name: Suffix:

Position/Title:

* Organization Name:

Department: Division:

* Street1:

Street2:

* City:

* Country: * ZIP / Postal Code:

* Phone Number: Fax Number:

* Email:

* State:

County / Parish:

Province:

Prefix:

Lochhead

Dr. Michael J

80303-1377

Boulder `

5603 Arapahoe Avenue,Suite 1

Vice President/CTO

MBio Diagnostics, Inc.

CO: Colorado

USA: UNITED STATES

	


 		

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SF 424 (R&R) APPLICATION FOR FEDERAL ASSISTANCE Page 2

15. ESTIMATED PROJECT FUNDING 16. * IS APPLICATION SUBJECT TO REVIEW BY STATE EXECUTIVE 

ORDER 12372 PROCESS?

a. YES a. Total Federal Funds Requested

17. By signing this application, I certify (1) to the statements contained in the list of certifications* and (2) that the statements herein are 

true, complete and accurate to the best of my knowledge. I also provide the required assurances * and agree to comply with any resulting 

terms if I accept an award. I am aware that any false, fictitious. or fraudulent statements or claims may subject me to criminal, civil, or 

administrative penalities. (U.S. Code, Title 18, Section 1001)

19. Authorized Representative

* First Name: Middle Name:

* Last Name: Suffix:

* Position/Title:

* Organization:

Department: Division:

* Street1:

Street2:

* City:

* State:

* Country: * ZIP / Postal Code:

* Phone Number: Fax Number:

* Email:

* Signature of Authorized Representative * Date Signed

20. Pre-application

* The list of certifications and assurances, or an Internet site where you may obtain this list, is contained in the announcement or agency specific instructions.

County / Parish:

c. Total Federal & Non-Federal Funds

18. SFLLL or other Explanatory Documentation

Province:

b. Total Non-Federal Funds

Prefix:

0.00

* I agree

Ms. Cynthia

Moore

Director of Finance and Administration

MBio Diagnostics, Inc.

5603 Arapahoe Avenue,Suite 1

Boulder `

CO: Colorado

USA: UNITED STATES 80303-1377

Add Attachment Delete Attachment View Attachment

Add Attachment Delete Attachment View Attachment

DATE:

THIS PREAPPLICATION/APPLICATION WAS MADE 

AVAILABLE TO THE STATE EXECUTIVE ORDER 12372 

PROCESS FOR REVIEW ON:

PROGRAM HAS NOT BEEN SELECTED BY STATE FOR 

REVIEW

PROGRAM IS NOT COVERED BY E.O. 12372; OR

Michael Lochhead 05/07/2013

b. NO

d. Estimated Program Income

	


 		

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424 R&R and PHS-398 Specific

Table Of Contents Page Numbers

SF 424 R&R Face Page------------------------------------------------------------------------------------------ 1

Table of Contents--------------------------------------------------------------------------------------------- 3

Performance Sites--------------------------------------------------------------------------------------------- 4

Research & Related Other Project Information------------------------------------------------------------------ 5

 Project Summary/Abstract (Description)---------------------------------------- 6

 Public Health Relevance Statement (Narrative attachment)---------------------------------------- 7

 Facilities & Other Resources---------------------------------------- 8

 Equipment---------------------------------------- 10

Research & Related Senior/Key Person-------------------------------------------------------------------------- 12

 Biographical Sketches for each listed Senior/Key Person---------------------------------------- 14

Research & Related Budget - Year 1---------------------------------------------------------------------------- 29

Research & Related Budget - Year 2---------------------------------------------------------------------------- 32

 Budget Justification---------------------------------------- 35

Research & Related Budget - Cumulative Budget----------------------------------------------------------------- 37

Research & Related Budget - Consortium Budget (Subaward 1)---------------------------------------------------- 38

SBIR/STTR Information----------------------------------------------------------------------------------------- 46

 Commercialization Plan---------------------------------------- 48

 Prior SBIR Phase II Awards---------------------------------------- 60

PHS 398 Specific Cover Page Supplement------------------------------------------------------------------------ 61

PHS 398 Specific Research Plan-------------------------------------------------------------------------------- 63

 Introduction---------------------------------------- 64

 Specific Aims---------------------------------------- 65

 Research Strategy---------------------------------------- 66

 Human Subjects Sections---------------------------------------- 78

 Protection of Human Subjects---------------------------------------- 78

 Women &Minorities---------------------------------------- 80

 Planned Enrollment Table---------------------------------------- 81

 Children---------------------------------------- 82

 Bibliography & References Cited---------------------------------------- 83

PHS 398 Checklist--------------------------------------------------------------------------------------------- 85

Table of Contents Page 3

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
County:

* ZIP / Postal Code:

* Country:

* State:

* City: 

Street2:

Organization Name:

Project/Performance Site Location(s)

Project/Performance Site Primary Location

* Street1:

Province:

MBio Diagnostics, Inc.

5603 Arapahoe Avenue, Suite 100

Boulder

CO: Colorado

USA: UNITED STATES

80303-1377

OMB Number: 4040-0010 

Expiration Date: 08/31/2011

I am submitting an application as an individual, and not on behalf of a company, state, 

local or tribal government, academia, or other type of organization. 

DUNS Number:

* Project/ Performance Site Congressional District: CO-002

Project/Performance Site Location 1

University of California, San Diego

220 Dickson Street, Suite A

San Diego

CA: California

USA: UNITED STATES

* ZIP / Postal Code: 92103-8950 CA-053

* Country:

Province:

* State:

* City: 

Street2:

* Street1:

Organization Name:

County:

DUNS Number:

* Project/ Performance Site Congressional District: 

I am submitting an application as an individual, and not on behalf of a company, state, 

local or tribal government, academia, or other type of organization. 

Additional Location(s) Add Attachment Delete Attachment View Attachment

	


 		

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Performance Sites Page 4

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
1. * Are Human Subjects Involved?

IRB Approval Date:

Human Subject Assurance Number:

2. * Are Vertebrate Animals Used?

IACUC Approval Date:

Animal Welfare Assurance Number

4.b. If yes, please explain:

4.c. If this project has an actual or potential impact on the environment, has an exemption been authorized or an environmental assessment (EA) or 

 environmental impact statement (EIS) been performed?

4.d. If yes, please explain:

6. * Does this project involve activities outside of the United States or partnerships with international collaborators?

6.b. Optional Explanation:

7. * Project Summary/Abstract

11. Equipment

8. * Project Narrative

12. Other Attachments

RESEARCH & RELATED Other Project Information

Is the IACUC review Pending?

If no, is the IRB review Pending?

2.a. If YES to Vertebrate Animals

3. * Is proprietary/privileged information included in the application? 

4.a. * Does this project have an actual or potential impact on the environment? 

6.a. If yes, identify countries:

1 2 3 4 5 6

9. Bibliography & References Cited

10. Facilities & Other Resources

Yes No

1.a If YES to Human Subjects

Yes No

Yes No

10/06/2011

If yes, check appropriate exemption number.

Yes No

Yes No

Yes No

Yes No

Yes No

Yes No

1239-Project Summary.pdf Add Attachment Delete Attachment View Attachment

1240-Project Narrative.pdf Add Attachment Delete Attachment View Attachment

1241-Bibliography.pdf Add Attachment Delete Attachment View Attachment

1242-MBio Facilities Other Resources Add Attachment Delete Attachment View Attachment

1243-MBio Equipment 20120823.pdf Add Attachment Delete Attachment View Attachment

Add Attachments Delete Attachments View Attachments

Is the Project Exempt from Federal regulations?

5. * Is the research performance site designated, or eligible to be designated, as a historic place? Yes No

5.a. If yes, please explain:

	


 		

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Other Information Page 5

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
MBio Diagnostics, Inc. proposes to develop a low-cost, point-of-care (POC), HIV-1/2 antigen/antibody 

combination diagnostic device. The goal is for the Ag/Ab combo system to deliver the performance of 

laboratory based, 4th generation antigen/antibody clinical analyzers, in a POC platform with cost and 

workflow of the widely used HIV rapid tests. Phase I research and development met or exceeded stated 

technical milestones. In particular, the MBio HIV antigen and antibody assays provided a combined 

performance that significantly exceeds performance FDA approved rapid tests, and provides near 

equivalence to much more complicated laboratory based tests. HIV infection remains a major public health 

crisis both in the United States and worldwide. There is increasing awareness that acutely infected 

individuals disproportionately contribute to disease spread. Yet these individuals remain the most difficult to 

identify, as infectivity is highest prior to the appearance of the HIV antibodies that serve as the basis for 

serological diagnostics. There are currently no FDA-approved point-of-care (POC) tests that directly target 

HIV viral antigens. An HIV-1 antigen/antibody (Ag/Ab) combination assay – the so-called “4th generation” 

immunoassay – in an inexpensive, simple to use, POC format would fundamentally improve HIV-1 

screening efforts in the United States and worldwide. The specific aims of this proposal are to: (1) Combine 

the Phase I p24 antigen detection assay with the MBio multiplexed serology assay cartridge, addressing 

issues of final monoclonal antibody (mAb) pair selection, HIV-1 Ag and HIV-2 Ag selection, reagent 

conjugations, cross-reactivity, and minimization of assay steps and complexity. (2) Modify the MBio 

Cartridge, Rack, Reader, and Software to deliver an automated HIV-1/2 Ag/Ab combo result, and 

incorporate heat stable assay reagents into the MQ cartridge. The Aim 2 milestone is a portable, integrated 

system delivered to clinical collaborators that meets FDA CLIA waiver guidance requirements. (3) Validate 

system using well characterized early HIV infection specimens including a panel of 200 HIV positive 

specimens comprised of 20 acute infection samples, early seroconversion, and seropositive samples. 200

HIV-negative samples will be used for specificity testing. (4) Place systems in an intended use setting and 

capture operational and usability feedback in advance of design lock, and to use this site to generate a 

preliminary dataset on capillary whole blood samples. The outcome of this program will be a system design 

and dataset for an FDA investigational device exemption (IDE) meeting in advance of clinical trials.

Project Description Page 6

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
The proposed point-of-care (POC) HIV-1 Ag/Ab combination diagnostic device will fill a critical unmet need 

for HIV screening technologies that detect recent and acutely infected individuals. The technology will be 

used in multi-test algorithms in public health laboratories, STD clinics, urban emergency departments, 

targeted outreach programs, etc., both in the United States and worldwide.

Public Health Relevance Statement Page 7

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
MBIO DIAGNOSTICS, INC. 

FACILITIES AND OTHER RESOURCES 

Biosafety and Biocontainment 

MBio operates a dedicated Biosafety Level 2 Laboratory that has been designed per guidance in the 

Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th Edition. MBio contracts certified 

industrial hygiene experts (Hellman and Associates, Wheat Ridge, CO) to perform annual training, safety 

audits, and written plan reviews. Per OSHA guidelines, MBio meets Bloodborne Pathogen training and 

vaccination requirements. Biological waste management is through a certified medical waste hauler 

(Colorado Medical Waste, Inc.). 

All MBio staff with exposure to human samples are also up-to-date on required Protection of Human 

Subjects training. 

General Facility Description 

MBio Diagnostics occupies a total of approximately 17,000 sq. ft. on Arapahoe in Boulder, CO with 

approximately 9,000 sq ft dedicated to laboratory and manufacturing. In addition to MBio’s technical staff 

members, the company has a 3 person G&A team that provides accounting, purchasing services, and 

human resources support, a 4 person clinical & regulatory team providing clinical, regulatory and quality 

support, and a 2 person marketing department. The MBio facility includes employee offices, cubicles, 

computer workstations and network support, conference rooms, projectors, etc. 

MBio’s laboratories are divided into 5 controlled access work areas, described here. 

Assay Wet Lab. The assay lab serves as the wet lab for standard assay development procedures. It is 

equipped with a sink, lab refrigerator, -20C freezer, dishwasher, sonicator and various mixers and shakers. 

Active glassware and lab disposables are stored here. The work space has lab benches for four scientists 

(~350 sq. ft). 

Biosafety Level 2 Laboratory. MBio operates a dedicated Biosafety Level 2 Laboratory that has been 

designed per guidance in the Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th Edition. 

The dedicated BL2 laboratory is a controlled access room housing a sink, three certified Class II biosafety 

cabinets, a state-of-the-art Beckman Allegra biosafety centrifuge. Refrigerators, a -20C freezer, and a -80C 

freezer are in the BL2 laboratory. 

PCR laboratories. The facility has a three-room PCR laboratory set up. The first of these laboratories is the 

pre-PCR reagent clean room. This is a 110 sq. ft. area dedicated to PCR reagent preparation and is 

equipped with a HEPA-filtered UV-equipped forced-air hood to prevent PCR contamination. The second 

PCR station resides in the BL2 laboratory with a dedicated biosafety cabinet to allow for safe nucleic acid 

extraction from biohazardous material and PCR target addition in a controlled environment. The final PCR 

station resides within the Assay wet lab. This lab houses the thermocycler, gel electrophoresis equipment 

and imaging systems and has sufficient space and resources for many downstream analyses. The three 

PCR stations are spatially separated, have been designed for unidirectional traffic flow, and each contain 

dedicated 4C refrigerators, -20C freezers, pipettes, and other common, necessary lab equipment. 

Facilities Page 8

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Instrument Laboratory. This ~190 sq. ft dedicated area houses the benches and optical tables used in 

instrument fluorescence waveguide instrument development. Test equipment, laboratory computers, and 

the fluorescence microscope are housed here. 

Array Fabrication Cleanroom. The ~200 sq. ft. cleanroom facility houses MBio’s Bio-Dot microarraying 

robot, enabling prototype waveguide substrate array printing in a cleanroom environment analogous to a 

full-scale production facility. 

A second fabrication room (~120 sq. ft.) focuses on final waveguide preparation and cartridge assem ly. 

This space includes a spin coater, pneumatic press, sonication baths used for cleaning, Sorvall centrifuge, a 

deionized water supply, and various scales and pumps.. 

 

Cartridge Development Area. A dedicated workspace has been created for disposable cartridge 

development. This area provides material storage, cutting tools, as well has peristaltic and syringe pump 

for controlled flowrate fluidic system development. Heat sealing equipment is available for cartridge 

packaging. Two Tenney environmental chambers, one with humidity control, are used for cartridge stability 

studies. The room also contains an AirClean Systems ductless chemical fume hood, benchtop vacuum 

ovens, and plasma treatment oven, used for surface chemistry preparations. The custom contact angle 

measurement tool in this room is used for surface characterization and quality control. 

Facilities Page 9

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Equipment

MBio Diagnostics operates a fully equipped R&D laboratory. Equipment is organized by location or 

affiliation to that location. The dedicated facilities listed here are described in detail in the Facilities 

document submitted with this grant.

Assay Wet Lab 

• Gel electrophoresis equipment (PAGE 

and agarose; prep and analytical)

• Bio-Rad Versadoc gel imager

• Bio-Rad GelDoc Imaging System

• -20°C Laboratory freezer (Kenmore) 

• Shaker (Barnstead/Labline) 

• Microfuge (VWR Galaxy Mini) 

• Various pipetters (Rainin LTS)

• Pipet-Aid automatic pipetters

• Hotplate-stirrers (VWR)

• Vortex mixers

• pH meters (Mettler)

• Analytical balance (A&D Limited) 

• Laboratory refrigerators (Kenmore, 

Frigidaire) 

• Microcentrifuges (Fisher accuSpin)

• Spectrophotometer (Thermo NanoDrop)

• Fisher Scientific IsoTemp temp block

• De-ionized and MilliQ water purification

• SPT Ice Maker

• PCR Thermal Cycler (Perkin Elmer 

2400)

• BiioRad CFX96 Real-Time Thermal 

Cycler System

• Luminex MagPix 

• BioCad 700E Perfusion 

Chromatography workstation

Cell Culture / Biosafety Level 2 Lab

• 3 Class II Biosafety Cabinets (4’ 

SterilGard, 6’ NuAire, 6’ Labconco) 

• CO2 Incubator (VWR 2350) 

• Centrifuge (Beckman Allegra X-12R) 

• Microplate Thermoshaker (Kisker 

Biotech PSF-100HL)

• Microplate reader (BioTek ELX800)

• Autoclave (Napco) 

• Laboratory refrigerator (VWR)

• -20°C Laboratory freezer (Frigidaire) 

• Cell Culture Plate Incubator

• Microcentrifuge (Fisher accuSpin)

• Water Bath (VWR)

• Pipetters

• Rotators and mixers

Pre-PCR

• PCR Workstation (Airclean 600)

• -20C freezer

• Refrigerator

• Pipetters

Common Area

• -80°C upright freezer (Forma Scientific) 

• -20C freezer (ScienTemp) 

• Liquid Nitrogen Freezer (dewar 

equipment with cryobox rack system)

• 2 Tenney Environmental chambers 

(T2RC and TUJR)

• Heat sealers for packaging

• 4 x Refrigerator

Instrument Lab

• Digital oscilloscope & multimeters

• Function generator

• CMOS cameras

• Lock-in amplifier

• Spectrum analyzers

• DC current/voltage supplies

• Laser diode drivers

• Laser power meter

• Lasers

• Various optics + mounts + hardware

• Various electronics

• Olympus IX-71 inverted fluorescence

microscope with phase optics and 

Retiga cooled CCD camera

Cleanroom/Chemistry Lab

• Ductless chemical fume hood, AirClean 

Systems XL10

• Vacuum ovens (VWR 1400E and VWR 

1430M ) 

• Labconco Filtermate auxiliary air filter

• Nitrogen purged drybox

• 2 x Plasma Oven (PlasmaEtch PE-200; 

Harrick Plasma PDC-001) 

• Vacuum Pumps (Edwards E2M8 and 

Edwards XDS-5 Dry Scroll)

Equipment Page 10

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
• Vacuum desiccator for process 

development

• Labconco Centrivap cold trap (# = 2)

• Custom Contact Angle Goniometer

• Bio-Dot AD3200 robotic arrayer 

equipped with Bio-Jet print head, 

humidity control system, and dedicated 

workstation with AxSys software 

package

Cartridge Assembly Area

• Pneumatic press (Sonitek TS500) 

• 2 x Spin Coater (CPK Industries; Laurell 

WS-650-23) 

• Swinging Bucket Centrifuge (Sorvall 

T6000D)

• Bath sonicators (Crest, # = 2)

• Stereomicroscope with video capture

• Syringe pump

• Peristalitic pump (Ecoline)

• Sink

• USFilter de-ionizer

• Denver Instruments P-402 Balance

• Symphony SB70P pH Meter

Equipment Page 11

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Province:

PROFILE - Project Director/Principal Investigator

Prefix: * First Name: Middle Name:

* Last Name: Suffix:

Organization Name: Division:

Position/Title: Department:

* Street1:

Street2:

* Phone Number: Fax Number:

* E-Mail:

Credential, e.g., agency login:

* Project Role: Other Project Role Category:

* Country: * Zip / Postal Code:

* State:

* City: County/ Parish:

Attach Current & Pending Support

RESEARCH & RELATED Senior/Key Person Profile (Expanded)

*Attach Biographical Sketch

Dr. Michael J

Lochhead

Vice President/CTO

MBio Diagnostics, Inc.

5603 Arapahoe Avenue,Suite 1

Boulder `

CO: Colorado

USA: UNITED STATES 80303-1377

PD/PI

1235-Lochhead_Biosketch_HIV-A Add Attachment Delete Attachment View Attachment

Add Attachment Delete Attachment View Attachment

OMB Number: 4040-0001 

Expiration Date: 06/30/2011

Degree Type:

Degree Year:

Province:

PROFILE - Senior/Key Person

Prefix: * First Name: Middle Name:

* Last Name: Suffix:

Organization Name: Division:

Position/Title: Department:

* Street1:

Street2:

* Phone Number: Fax Number:

* E-Mail:

Credential, e.g., agency login:

* Country: * Zip / Postal Code:

* State:

* City: County/ Parish:

Dr.

Senior Scientist

MBio Diagnostics, Inc.

USA: UNITED STATES

Daniel 

Nieuwlandt

5603 Arapahoe Avenue, Suite 1

Boulder

CO: Colorado

80303-1377

* Project Role: Other (Specify) Other Project Role Category: Senior Scientistg

1

Degree Type:

Degree Year:

*Attach Biographical Sketch 1236-Nieuwlandt_Biosketch_P22 Add Attachment Delete Attachment View Attachment

Attach Current & Pending Support Add Attachment Delete Attachment View Attachment

	


 		

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Key Personnel Page 12

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED Senior/Key Person Profile (Expanded)

Province:

PROFILE - Senior/Key Person

Prefix: * First Name: Middle Name:

* Last Name: Suffix:

Organization Name: Division:

Position/Title: Department:

* Street1:

Street2:

* Phone Number: Fax Number:

* E-Mail:

Credential, e.g., agency login:

* Country: * Zip / Postal Code:

* State:

* City: County/ Parish:

Dr.

Professor of Medicine

University of California, San Diego

USA: UNITED STATES

Constance

Benson

200 W. Arbor Drive, MC#8208

San Diego

CA: California

82103-8960

* Project Role: Co-Investigator Other Project Role Category:

2

Degree Type:

Degree Year:

*Attach Biographical Sketch 1237-Benson BioSketch.pdf Add Attachment Delete Attachment View Attachment

Attach Current & Pending Support Add Attachment Delete Attachment View Attachment

Province:

PROFILE - Senior/Key Person

Prefix: * First Name: Middle Name:

* Last Name: Suffix:

Organization Name: Division:

Position/Title: Department:

* Street1:

Street2:

* Phone Number: Fax Number:

* E-Mail:

Credential, e.g., agency login:

* Country: * Zip / Postal Code:

* State:

* City: County/ Parish:

Dr. J

Professor of Medicine

University of California, San Diego

USA: UNITED STATES

Susan

Little

200 W. Arbor Drive, MC8208

San Diego

CA: California

92103-8950

* Project Role: Co-Investigator Other Project Role Category:

3

Degree Type:

Degree Year:

*Attach Biographical Sketch 1238-Little Biosketch.pdf Add Attachment Delete Attachment View Attachment

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Key Personnel Page 13

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
BIOGRAPHICAL SKETCH

Provide the following information for the Senior/key personnel and other significant contributors.

Follow this format for each person. DO NOT EXCEED FOUR PAGES.

NAME

Michael Joseph Lochhead

POSITION TITLE

Vice President, MBio Diagnostics, Inc.

eRA COMMONS USER NAME (credential, e.g., agency login)

EDUCATION/TRAINING (Begin with baccalaureate or other initial professional education, such as nursing, include postdoctoral training and 

residency training if applicable.)

INSTITUTION AND LOCATION DEGREE

(if applicable) MM/YY FIELD OF STUDY

University of Notre Dame, Notre Dame, IN B.S. 1988 Chemical Engineering

University of Notre Dame, Notre Dame, IN B.A. 1988 Government

University of Wisconsin, Madison, WI Ph.D. 1995 Chemical Engineering

University of Washington, Seattle, WA Post-Doc 1996-1998 Bioengineering

A. Personal Statement 

The purpose of this project is to integrate the quantitative p24 direct antigen detection assay into the MBio 

POC diagnostics product development path. I have the expertise and leadership required to carry out the 

proposed work in a cost effective manner. Over the last 5 years, I have managed growth of the MBio 

Diagnostics from a 2 person R&D group within Precision Photonics Corporation (Boulder, CO) to a medical 

diagnostics company incorporated in early 2010. As Vice President since the Company’s inception, I lead 

research, development, and commercialization efforts for MBio’s product pipeline. I am Principal Investigator 

on multiple major contracts with the National Institutes of Health, and I manage clinical collaborations in the 

United States, Brazil, Kenya, and Mozambique. While at Accelr8 Technology Corp. (Denver, CO), I 

successfully moved two surface chemistry products targeted specifically for microarray applications from 

conception through R&D and commercialization. While at Accelr8 I was also project manager for the 

company’s prototype microscopy-based microbiology instrument, which combined integrated plastic 

microfluidic disposables and a fluidic control system. My academic career included an Assistant Professor 

position in the Department of Chemical Engineering at the University of New Hampshire where my group 

focused on advanced materials for biosensor and biomaterial applications. This followed my post-doctoral 

training in the Bioengineering program at the University of Washington where my research focuses included 

biomaterials engineering and basic science and microfluidic device development. I am co-inventor on multiple 

issued and pending patents. In summary, I have a demonstrated record of successful project management in 

the areas of assay devices, surface chemistry, and instrumentation, all of which are relevant to the current 

project. 

B. Positions and Honors 

Positions and Employment

1988-1990 Staff Engineer, ICF Incorporated, Fairfax, VA 

1990-1995 Graduate Studies, Department of Chemical Engineering, University of Wisconsin, Madison, WI, 

1996-1997 Post-Doctoral Research Associate, Department of Bioengineering, University of Washington, 

Seattle, WA 

1998-2001 Assistant Professor, Department of Chemical Engineering, University of New Hampshire, 

Durham, NH 

2001-2007 Senior Scientist, Accelr8 Technology Corporation, Denver, CO

2007-2008 Diagnostics Team Leader, Precision Photonics Corporation, Boulder, CO 

2008-2010 Vice President, MBio Diagnostics division of Precision Photonics Corporation, Boulder, CO 

2010-present Vice President, MBio Diagnostics, Inc., Boulder, CO 

Biosketches Page 14

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Other Experience and Professional Memberships: 

1999-2001 Participating Faculty, Center to Advance Molecular Interaction Sciences (CAMIS), University of 

New Hampshire (1999-2001) 

1995-2007 Member, American Chemical Society 

2008- Member, American Association for Clinical Chemistry 

2008- Journal reviewer for Analytical Chemistry (American Chemical Society) 

2009- Journal reviewer for Lab on Chip (Royal Society of Chemistry) 

2009- Journal reviewer for Physical Chemistry Chemical Physics (Royal Society of Chemistry) 

2009- Journal reviewer for Analyst (Royal Society of Chemistry) 

2010- NIH National Cancer Institute, SBIR Program Peer Review Committee 

Honors: 

1993 Ragatz Outstanding Teaching Award, University of Wisconsin, Madison, WI 

1993-1994 Amoco Research Fellowship, Chemical Engineering, University of Wisconsin, Madison, WI 

1993-1995 Teaching Assistant Fellow, College of Engineering, University of Wisconsin, Madison, WI 

C. Selected Peer-reviewed Publications 

1. Lochhead, M. J.; Letellier, S. R.; Vogel, V., Assessing interfacial electrostatics in oriented mineral 

nucleation at ordered organic monolayers. J Phys Chem B 1997, 101, 10821. 

2. Letellier, S. R.; Lochhead, M. J.; Campbell, A. A.; Vogel, V., Oriented growth of calcium oxalate 

monohydrate crystals beneath phospholipid monolayers. Biochim Biophys Acta 1998, 1380, (1), 31-45. 

3. Lochhead, M. J.; Touryan, L. A.; Vogel, V., In situ analysis of europium-calcium-oxalate crystallization 

using luminescence microspectroscopy. J Phys Chem B 1999, 103, 3411. 

4. Clarner, M. A.; Lochhead, M. J., Hybrid Micro-Optical Sensors via Sol-Gel Soft Lithography. Mat. Res. 

Soc. Symp. Ser. 2000, 628, CC9.3.1-CC9.3.6. 

5. Metzger, S.; Lochhead, M. J.; Grainger, D. W., Surface technologies to improve performance in protein 

microarray based molecular diagnostics. IVD Technology 2002, 8, (5), 39-45. 

6. Touryan, L. A.; Lochhead, M. J.; Marquardt, B. J.; Vogel, V., Sequential switch of biomineral crystal 

morphology using trivalent ions. Nat Mater 2004, 3, (4), 239-43. 

7. Grainger, D. W.; Greef C. H.; Gong, P.; Lochhead, M. J., Current Microarray Surface Chemistries, in 

Microarrays, Second Edition, Volume I: Synthesis Methods, J. Rampal, editor; Humana Press, Methods in 

Molecular Biology series, 2007. 

8. Saldarriaga Fernandez, I.C; van der Mei, H. C.; Lochhead, M. J.; Grainger, D. W.; Busscher, H. J., 

Bacterial adhesion onto surfaces of a multi-component cross-linked poly(ethylene glycol)-based polymer 

coating. Biomaterials 2007, 28, 4105-4112. 

9. Harbers, G. M.; Emoto, K.; Greef, C.; Metzger, S. W.; Woodward, H. W.; Mascali, J. J.; Grainger, D. W.; 

Lochhead, M. J., Functionalized Poly(ethylene glycol)-based Bioassay Surface Chemistry that Facilitates 

Bio-Immobilization and Inhibits Nonspecific Protein, Bacterial, and Mammalian Cell Adhesion. Chem. Mat.

2007, 19(18), 4405-4414. 

10. Myatt, C.J.; Delaney, M.J.; Todorof, K.; Heil, J.R.; Givens, M.; Schooley, R.T.; Lochhead, M.J, Low-cost, 

multiplexed biosensor for disease diagnosis, Proc. SPIE 7167 Frontiers in Pathogen Detection: From 

Nanosensors to Systems, 716703 (2009) 

11. Lochhead, M.J., Insights from the 2010 HIV Diagnostics Conference. Expert Rev Mol Diagn 2010, 10(5) 

565-567. 

12. Lochhead M.J., Todorof K., Delaney M., Ives J.T., Greef C., Moll K., et al. Rapid Multiplexed 

Immunoassay for Simultaneous Serodiagnosis of HIV-1 and Co-Infections. J Clin Microbiol. 2011, 49(10) 

3584-3590.

13. Devlin S, Meneely JP, Greer B, Greef C, Lochhead MJ, Elliott CT. Next generation planar waveguide 

detection of microcystins in freshwater and cyanobacterial extracts, utilising a novel lysis method for 

portable sample preparation and analysis. Anal Chim Acta. 2013; 769:108-13. 

14. Meneely JP, Campbell K, Greef C, Lochhead MJ, Elliott CT. Development and validation of an 

ultrasensitive fluorescence planar waveguide biosensor for the detection of paralytic shellfish toxins in 

marine algae. Biosens Bioelectron. 2013; 41:691-7.

Biosketches Page 15

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
15. Logan C, Givens M, Ives JT, Delaney M, Lochhead MJ, Schooley RT, et al. Performance evaluation of the 

MBio Diagnostics point-of-care CD4 counter. J Immunol Methods. 2013; 387(1-2):107-13. PMCID: 

3529779. 

D. Research Support 

Ongoing Research Support: 

NIH NIAID 5R44AI070052-05 Lochhead (PI) 03/01/2006 – 02/28/2014 

Low Cost Laser Diagnostic for CD4+ T Cell Counting 

The goal of this project is to build on the cell enumeration instrument developed under NIST-ATP FY2007 

70NANB7H7053, improving count statistics, adding additional CD markers, and performing larger volume 

clinical sample testing. 

Role: Principal Investigator 

NIH NIAID 5R01AI096189-02 Lochhead (PI) 07/05/2011 – 06/30/2016 

Integrated System for Rapid Detection of Respiratory Pathogens 

The goal of this project is development of a low-cost, no amplification, nucleic acid diagnostic technology for 

respiratory pathogens. 

Role: Principal Investigator 

NIH NIAID 2R44AI68543-06 Myatt (PI) 09/01/2009 – 08/31/2013 

Development of Low Cost Multi-Pathogen Laser Diagnostics for HIV and AIDS Co-Infections 

The goal of this project is development of an inexpensive, multiplexed serotyping and antigen detection pointof-care diagnostics platform, targeted at HIV and opportunistic infection screening. 

Role: Project Manager, Scientist 

Completed Research Support (last 3 years)

NIH U54 AI065359 Barbour (PI) 05/01/2009 – 04/30/2011 

Subaward No. 2009-2167 Myatt (Subaward, PI) 

Pacific Southwest Regional Center of Excellence for Biodefense and Emerging Infectious Diseases. Project 

2.5: Benchtop Devices for Detection of BoNT in Clinical Samples 

The goal of this project is development of a clinical assay system based on the botulinum toxin Assay with 

Large Immono-Sorbent Surface Area (ALISSA) format. The aim of the subaward project is to create, test, and 

validate waveguide-based detection system that will be integrated with the automated ALISSA instrument. 

Role: Project Manager, Scientist 

NIH NIAID 1R43AI093289-01A1 Lochhead (PI) 08/10/2011 – 07/31/2012 

Point-of-Care HIV Antigen/Antibody Diagnostic Device 

The goal of this project is to develop a low-cost, point-of-care (POC), HIV-1 antigen/antibody combination 

diagnostic device using MBio’s novel multiple particle assay approach. The system goal is to deliver 

performance of laboratory based, clinical analyzers, in a POC platform with cost and workflow of widely used 

HIV rapid tests. 

NIH NIAID 7R43AI091097-02 Lochhead (PI) 07/03/2010 – 01/31/2011 

Multiple Antigen Point-of-Care Device for Improved Syphilis Diagnosis 

The goal of this project is development of a quantitative non-treponemal assay to complement mBio’s 

treponemal-specific rapid diagnostic. The combined treponemal / non-treponemal test will significantly 

advance point-of-care syphilis testing. 

Role: Principal Investigator 

NIH NIAID 1U01AI074521-04 Schooley (PI) 07/01/2007 – 06/30/2012 

Multiplex Nucleic Acid Detection Device for the Diagnosis of Respiratory Viruses 

Biosketches Page 16

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
The goal of this project is development of the low-cost, field deployable instrumentation and analysis platform 

for multiplexed nucleic acid assays, targeting influenza virus subtyping specifically and respiratory pathogen 

identification in general. 

Role: Project Manager, Scientist 

Biosketches Page 17

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
BIOGRAPHICAL SKETCH

Provide the following information for the Senior/key personnel and other significant contributors. 

Follow this format for each person. DO NOT EXCEED FOUR PAGES.

NAME

Daniel Terry Nieuwlandt

POSITION TITLE

Senior Scientist, MBio Diagnostics, Inc.

eRA COMMONS USER NAME (credential, e.g., agency login)

EDUCATION/TRAINING (Begin with baccalaureate or other initial professional education, such as nursing, include postdoctoral training and 

residency training if applicable.)

INSTITUTION AND LOCATION DEGREE

(if applicable) MM/YY FIELD OF STUDY

California State University, Fullerton, CA B.A. 1985 Biological Science

Iowa State University, Ames, IA M.S. 1988 Microbiology

Ohio State University, Columbus, OH Ph.D. 1992 Microbiology

University of Colorado, Boulder, CO Post. Doc. 1992-1994 Molecular Biology

A. Personal Statement

With over 25 years of research experience, including 18 years with a primary focus on assay development at 

multiple early-phase biotechnology companies, I bring a breadth of research and project management 

experience to this project. Of particular relevance is my extensive experience with IVD development and 

bioconjugation techniques. As a Senior Scientist immersed in HIV and influenza IVD R&D at MBio 

Diagnostics, I have extensive experience with all aspects of the company’s planar waveguide technology. 

As Director of Assay Development at Lyzer Diagnostics, Inc., I brought an "Enhanced Velocity 

Electroimmunoassay" platform (high sensitivity solution phase immunoassay) from concept to a functioning 

prototype. At Great Basin Scientific, I led a team of scientists that developed a rapid automated molecular 

assay for Staphylococcal species. As an assay development Group Leader at SomaLogic, Inc., I led the 

successful development of sensitive and highly specific nucleic acid-based microarray diagnostic assays 

based on the company's photoaptamer microarray technology. I additionally held the positions of Director of 

Biology and Vice President, Biology, at Invenux, Inc., and Associate Research Director at NeXstar 

Pharmaceuticals, Inc. At Invenux, I directed a project aimed at discovering antibiotics active against MRSA, 

a research effort that required the cloning, expression, and purification of PBP2a and amino acid adding 

enzymes (peptidoglycan biosynthesis pathway) and the development of assays for enzyme activity and 

methods for catalytic RNA partitioning. My role at NeXstar included the development of aptamer reagents 

specific for Group A Streptococci and Chlamydia elementary bodies and the management of a collaborative 

diagnostics research effort with Becton Dickinson. Each of the research and development efforts at MBio 

Diagnostics, Lyzer Diagnositcs, Somalogic, Invenux, and NeXstar required the conjugation of proteins or 

nucleic acids to dyes and microspheres and the use of these constructs as IVD reagents. I am a co-inventor 

on six issued patents. In brief, I am a microbiologist with extensive molecular biology experience and a track 

record of successful IVD development.

B. Positions and Honors

Positions and Employment

1992-1994 Postdoctoral Research Fellow; University of Colorado, Boulder, CO

1994-1996 Senior Scientist; NeXagen, Inc. / NeXstar Pharmaceuticals, Inc., Boulder, CO

1996-1999 Associate Director; NeXstar Pharmaceuticals, Inc., Boulder, CO

2000-2001 Director of Biology; Invenux, Inc., Denver, CO

2001-2003 Vice President of Biology; Invenux, Inc., Denver, CO

2003-2008 Scientific Advisory Board Member; Cropsolution, Inc.

2004-2007 Senior Scientist & Assay Development Group Leader; Somalogic, Inc., Boulder, CO

2007-2008 Director of Assay Development; Lyzer Diagnostics, Inc. Boulder, CO

2008-2010 Director of Research; Great Basin Scientific, Longmont, CO

2010-2010 Director of Assay Development (Jan-Aug); Lyzer Diagnostics, Inc. Boulder, CO

2010-present Senior Scientist; MBio Diagnostics, Inc., Boulder, CO

Biographical Sketches for each listed Senior/Key Person 2 Page 18

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Honors

1987 Gamma Sigma Delta Honor Society of Agriculture membership.

1991 Phi Kappa Phi Honor Society membership.

1991 Proctor and Gamble Research Fellowship for “Transfer RNA Processing in the 

 Archaebacterium Haloferax volcanii."

1991 Graduate Student Alumni Research Award from Ohio State University for 

“Characterization of Ribonuclease P from the Archaebacterium Haloferax volcanii."

C. Peer-Reviewed Publications

1. Nieuwlandt, D. T., and P. A. Pattee. 1989. Transformation of a conditional peptidoglycan-deficient 

mutant of Staphylococcus aureus with plasmid DNA. J. Bacteriol. 171: 4906-4913.

2. Thompson, L. D., L. D. Brandon, D. T. Nieuwlandt, and C. J. Daniels. 1989. Transfer RNA intron 

processing in the halophilic archaebacteria. Can. J. Microbiol. 35: 36-42.

3. Nieuwlandt, D. T., and C. J. Daniels. 1990. An expression vector for the archaebacterium Haloferax 

volcanii. J. Bacteriol. 172: 7104-7110.

4. Nieuwlandt, D. T., E. S. Haas, and C. J. Daniels. 1991. The RNA component of RNase P from the 

archaebacterium Haloferax volcanii. J. Biol. Chem. 266: 5689-5695.

5. Nieuwlandt, D.T., M. B. Carr, and C. J. Daniels. 1993. In vivo processing of an intron-containing archael 

tRNA. Molec. Microbiol. 8: 93-99.

6. Palmer, J. R., D. T. Nieuwlandt, and C. J. Daniels. 1994. Expression of a yeast intron-containing tRNA 

in the archaeon Haloferax volcanii. J. Bacteriol. 176: 3820-3823.

7. Nieuwlandt, D. T., M. Wecker, and L. Gold. 1995. In vitro selection of RNA ligands to substance P. 

Biochemistry 34: 5651-5659.

8. Lin, Y., D. Nieuwlandt, A. Megallanez, B. Feistner, and S. D. Jayasena. 1996. High-affinity and specific 

recognition of human thyroid stimulating hormone (hTSH) by in vitro-selected 2’-amino-modified RNA. 

Nucleic Acids Res. 24: 3407-3414.

9. Nieuwlandt, D. 1998. In vitro selection of functional nucleic acid sequences. Pp. 117-132 In: Genetic 

Engineering with PCR. R. M. Horton and R. C. Tait, eds. Horizon Scientific Press. 

Portland, Oregon.

10. Nieuwlandt, D. 2000. In vitro selection of functional nucleic acid sequences. Curr. Issues Mol. Biol. 2: 

9-16.

11. Dewey, T. M., D. Nieuwlandt, and T. Tarasow. 2002. Integrated drug discovery technology in a test 

tube. Current Drug Discovery. July 2002: 21-25.

12. Nieuwlandt, D., M. West, X. Cheng, G. Kirshenheuter, and B. E. Eaton. 2003. The first example of an 

RNA urea synthase: selection through the enzyme active site of human neutrophile elastase. 

Chembiochem. 4: 651-654.

13. Tarasow, T.M., E. Kellogg, B.L. Holley, D. Nieuwlandt, S.L. Tarasow, and B.E. Eaton. 2004. The effect 

of mutation on RNA Diels-Alderases. J. Am. Chem. Soc. 126(38): 11843-51.

14. Gold, L., D. Ayers, J. Bertino, C. Bock, A. Bock, E.N. Brody, J. Carter, et al. 2010. Aptamer-based 

multiplexed proteomic technology for biomarker discovery. PLoS One 5(12): e15004.

15. Pasko, C., B. hicke, J. Dunn, H. Jaeckel, D. Nieuwlandt, D. Weed, E. Woodruff, X. Zheng, and R. 

Jenison. 2012. Staph ID/R: a rapid method for determining staphylococcus species identity and detecting 

the mecA gene directly from positive blood culture. J. Clin. Microbiol. 50(3): 810-7.

D. Research Support

Ongoing Research Support: 

NIH NIAID 5R01AI096189-02 Lochhead (PI) 07/05/2011 – 06/30/2016

Integrated System for Rapid Detection of Respiratory Pathogens

The goal of this project is development of a low-cost, no amplification, nucleic acid diagnostic technology for 

respiratory pathogens. 

Role: Senior Scientist

Biographical Sketches for each listed Senior/Key Person 2 Page 19

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Biographical Sketches for each listed Senior/Key Person 2 Page 20

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Biographical Sketches for each listed Senior/Key Person 3 Page 21

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J

BIOGRAPHICAL SKETCH 

Provide the following information for the Senior/key personnel and other significant contributors. 

Follow this format for each person. DO NOT EXCEED FOUR PAGES. 

NAME POSITION TITLE 

Constance A. Benson, M.D. 

_e_R_A_c_o_M_M_O_N_s_u_s_E_R_N_A_M_E ___________ _, Professor of Medicine 

EDUCATION/TRAINING (Begin with baccalaureate or other initial professional education, such as nursing, and include postdoctoral training and 

residency training if applicable . .) 

INSTITUTION AND LOCATION DEGREE YEAR(s) FIELD OF STUDY (if applicable) 

The Ohio State University, Columbus, OH B.A. 1975 Zoology 

The Ohio State U. College of Med., Columbus, OH M.D. 1978 Medicine 

Rush University Medical Center (Chicago, IL) Residency 1979-81 Internal Medicine 

Rush University Medical Center (Chicago, IL) Fellowship 1986 Infectious Diseases 

A. Personal Statement 

My role on the project will be to participate as the UCSD collaborative partner Pl, and to participate in and 

oversee the design, development, implementation and conduct of the research at UCSD; to facilitate and 

oversee the recruitment, monitoring and followup of research participants; to oversee site and laboratory 

acquisition of samples and conduct of study procedures by research staff; to oversee sample and data 

collection and submission; and to participate in the analysis and interpretation of data and publication of results 

as appropriate. I am highly qualified for this role based on my experience as an internationally recognized 

translational and clinical researcher in the field of HIV/AIDS, serving in the investigative and scientific 

leadership of the NIH/NIAID AIDS Clinical Trials Group (ACTG) continuously from 1996 to 2010 (as Pl/Chair 

from 2003-2010), and in these roles managing this multinational clinical trials network comprising 72 clinical 

research sites and laboratories in 14 different countries including the U.S., Haiti, Brazil, Peru, South Africa, 

Uganda, Zambia, Botswana, Malawi, Zimbabwe, Tanzania, Kenya, India, and Thailand. My responsibilities in 

the leadership of the ACTG included establishing and mentoring the development of investigators and 

research staff at 22 international clinical research sites in resource-limited settings and training more than 200 

international investigators, affiliated research and laboratory personnel at these sites to conduct scientifically 

rigorous randomized clinical trials and laboratory research. In the context of these activities, I have overseen 

the establishment, training and development of four major reference laboratories to conduct sophisticated and 

quality controlled viral genotypic resistance testing and sophisticated diagnostic testing for tuberculosis (TB) in 

support of the ACTG Network trials. My own research experience has included development of new drugs and 

strategies for treatment of HIV-1 infection, HIV-related opportunistic infections, and for the treatment and 

prevention of TB and other mycobacterial infections. I have chaired numerous multicenter clinical trials 

including serving as protocol chair, vice chair or co-investigator for 12 current ACTG clinical trials related to 

treatment or prevention of HIV and mycobacterial infections. In addition to these research roles, I am a 

member of the NIH/NIAID/DAIDS TB Laboratory Working Group; a member of the WHO Stop TB/HIV Working 

Group; a member of the ACTG's TB Transformative Science Group; a co-investigator in another SBIR grant to 

develop a point-of-care diagnostic instrument and assays for rapid quantitation of CD4 cell count and 

serodiagnosis of other infectious diseases; and a member of the External Scientific Advisory Group for the 

NIAID/DMID's TB Clinical Diagnostics Research Consortium. 

B. Positions and Honors 

Professional and Academic Appointments 

1981-1984 U.S. Navy Staff Physician, Great Lakes Naval Regional Medical Center; North Chicago, IL 

1986-1987 Instructor in Medicine, Rush Medical College/Rush-Presbyterian-St. Luke's Medical Center 

1987-1992 Assistant Professor of Medicine, Rush Medical College; Chicago, IL 

1993-1997 Associate Professor of Medicine, Rush Medical College; Chicago, IL 

1997-2004 Professor of Medicine, University of Colorado Health Sciences Center; Denver, CO 

2004-Present Professor of Medicine, University of California, San Diego; San Diego, CA 

2006-Present Infectious Diseases Training Program Director, UCSD, San Diego, CA 
Biographical Sketches for each listed Senior/Key Person 3 Page 22

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J

Administrative and Committee Positions 

1994-Present Co-Chair, USPHS/IDSA Treatment and Prevention of Opportunistic Infections Working Group 

1994-Present Member, Scientific Program Committee, Conference on Retroviruses and Opportunistic 

1995-2002 

1995-2003 

1996-Present 

2000-2003 

2003-2010 

Infections (Vice Chair 1997-1999; Chair 2000-2002) 

Vice Chair, Adult AIDS Clinical Trials Group, NIAID/NIH 

Chair, Scientific Agenda Steering Committee, Adult AIDS Clinical Trials Group, NIAID/NIH 

Member, Board of Directors, International AIDS Society - USA 

Member, NIH Office of AIDS Research Advisory Council (OARAC) (Chair 2001-2003) 

Chair and Principal Investigator, AIDS Clinical Trials Group network, NIAID/NIH 

2006-Present Director, UCSD Antiviral Research Center, San Diego, CA 

C. Selected peer-reviewed publications relevant to this project (from over 140, in chronological order). 

1. Benson CA, Williams PL, Currier JS, et al. A prospective, randomized trial examining the efficacy and 

safety of clarithromycin in combination with ethambutol, rifabutin, or both for the treatment of disseminated 

Mycobacterium avium complex disease in persons with acquired immunodeficiency syndrome. Clin Infect 

Dis 2003; 37(9):1234-1243. 

2. Benson CA, Kaplan JE, Masur H, Pau A, Holmes KK. Treating opportunistic infections among HIVinfected adults and adolescents: recommendations from CDC, the National Institutes of Health, and the 

HIV Medicine Association/Infectious Diseases Society of America. MMWR 2004; 53(RR-15):1-112. (also 

published in Clin Infect Dis 2005) 

3. Benson CA, Vaida F, Havlir DV, et al. A randomized trial of treatment interruption before optimized 

antiretroviral therapy for persons with drug-resistant HIV: 48-week virologic results of ACTG A5086. J 

Infect Dis 2006; 194(9):1309-1318. 

4. Bosch RJ, Bennett K, Collier AC, Zackin R, Benson CA. Pre-treatment factors associated with 3-year 

(144-week) virologic and immunologic responses to potent antiretroviral therapy. J Acquir Immune Defic 

Syndr 2007; 44(3):268-277. 

5. Smurzynski M, Collier AC, Koletar SL, Bosch RJ, Wu K, Bastow B, Benson CA. AIDS Clinical Trials Group 

Longitudinal Linked Randomized Trials (ALLRT): rationale, design, and baseline characteristics. HIV Clin 

Trials 2008; 9(4):269-282. 

6. Kitahata MM, Gange SJ, Abraham AG, Merriman B, ... Benson CA, et al. Effect of early versus deferred 

antiretroviral therapy for HIV on survival. N Engl J Med 2009; 360(18): 1815-26. 

7. Kumarasamy N, Madhavan V, Venkatesh KK, Saravanan S, Kantor R, Balakrishnan P, Devaleenal B, 

Poongulali S, Yepthomi T, Solomon S, Mayer KH, Benson C, Schooley R. High frequency of clinically 

significant mutations after first-line generic highly active antiretroviral therapy failure: implications for 

second-line options in resource-limited settings. Clin Infect Dis 2009; 49(2):306-9. 

8. Smith OM, May SJ, Perez-Santiago J, Strain MC, Ignacio CC, Haubrich RH, Richman DD, Benson CA, 

Little SJ. The use of pooled viral load testing to identify antiretroviral treatment failure. AIDS 2009; 23:2151-

8. Park D, Qin H, Jain S, Preziosi M, Minuto JJ, Mathews WC, Moser KS, Benson CA. Tuberculosis due to 

Mycobacterium bovis in patients co-infected with HIV. Clin Infect Dis 201 O; 51: 1343-6. 

9. Ribaudo HJ, Benson CA, Zheng Y, Koletar SL, Collier AC, Lok JJ, Smurzynski M, Bosch RJ, Bastow B, 

Schouten JT, for the ACTG A5001/ALLRT Protocol Team. No immediate and long-term risk of mycocardial 

infarction associated with initial antiretroviral treatment containing abacavir: results from ACTG 

A5001/ALLRT over 1 and 6 years. Clin Infect Dis 2011; 52:929-40. 

10. Krishnan S, Schouten JT, Jacobson DL, Benson CA, Collier AC, Koletar SL, Mitsuyasu R, for the ACTGALLRT Protocol Team. Incidence of non-AIDS defining cancer (NADC) in antiretroviral treatment (ART) 

na"ive subjects after ART initiation: an ALLRT (ACTG Longitudinal Linked Randomized Trials) analysis. 

Oncology 2011; 80:42-49. 

11. Daar ES, Tierney C, Fischl MA, Sax PE, Mollan D, Budhathoki C, Godfrey C, Jahed NC, Myers L, 

Katzenstein D, Farajallah A, Rooney JF, Pappa KA, Woodward WC, Patterson K, Bolivar H, Benson CA, 

Collier AC, for the AIDS Clinical Trials Group Study A5202 Team. Atazanavir plus ritonavir or efavirenz as 

part of a three drug regimen for initial treatment of HIV-1: a randomized trial. Ann Intern Med 2011; 

154:445-56. 

12. Krishnan S Wu K, Smurzynski M, Bosch RJ, Benson CA, Collier AC, Klebert MK, Feinberg J, Koletar SL, 

for the ALLRT/A5001 Team. Incidence rate of and factors associated with loss-to-follow-up in a longitudinal 

cohort of anti-retroviral treated HIV-infected persons: an AIDS Clinical Trials Group (ACTG) Longitudinal 

Linked Randomized Trials (ALLRT) analysis. HIV Clin Trials 2012; (in press). 
Biographical Sketches for each listed Senior/Key Person 3 Page 23

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J

13. Lochhead MJ, Todorof K, Delaney M, Ives JT, Greef C, Moll K, Rowley K, Vogel K, Myatt C, Zhang X-Q, 

Benson C, Reed S, Schooley RT. Rapid multiplexed immunoassay for simultaneous serodiagnosis of HIV1 and co-infections. J Clin Microbiol 2011; (Epub ahead of print] (in press). 

14. Eastburn A, Scherzer R, Zolopa AR, Benson C, Russell T, Do T, Bacchetti P, Shlipak M, Grunfeld C, Tien 

PC. Association of low-level viremia with inflammation and mortality in HIV-infected adults. PLoS ONE 

2011; 6(11 ):e26320. 

15. Thompson MA, Aberg JA, Hoy JF, Telenti A, Benson C, Cahn P, et al. Antiretroviral treatment of adult HIV 

infection: 2012 recommendations of the International Antiviral Society-USA panel. JAMA 2012; 308:387-

402. 

D. Research Support 

Active: 

5 UM1 AI069432 (Benson, Constance) 02/01/2007-11/30/2013 

National Institutes of Health 

UCSD Department of Medicine HIV/AIDS Clinical Trials Unit 

Major goals: The UCSD Department of Medicine HIV/AIDS Clinical Trials Unit conducts investigator-initiated 

clinical trials of the NIH/NIAID Adult AIDS Clinical Trials Group (AACTG), intended to develop new or improve 

existing antiviral therapies or strategies for the treatment and prevention of HIV infection and its complications 

in adults and older adolescents infected with HIV. (Role: Pl) 

7 UM1 Al68636 (Kuritzkes) 06/29/2006-5/31/2013 

NIH/Social and Scientific Systems, Inc. 

AIDS Clinical Trials Group Network Leadership Group 

Major goals: This is the Central Group Core for the AIDS Clinical Trials Group (AACTG), a large international 

multicenter clinical trials network that conducts investigator initiatied therapeutic clinical trials to develop new or 

improve existing antiviral drugs and strategies for the treatment and prevention of HIV infection and its 

complications. Dr. Benson serves as the immediate past Principal Investigator of the U01 and a current 

member of the Executive Committee of this Group. (Role: Co-Investigator) 

5 P01 AI074621 (Little, Susan) 

NIH/NIAID 

Determinants of HIV Transmission 

03/15/2008-02/28/2013 

Major goals: To focus on the identification and systematic evaluation of individuals who have been recently 

infected with HIV and the sexual partners who transmitted HIV to them (Transmission Pairs) to elucidate and 

quantify epidemiologic, behavioral, biologic, virologic, and host factors that contribute to transmission. (Role: 

Co-Investigator) 

Social & Scientific Systems, Inc (Gilbert, Tari) 07/02/2007-06/30/2013 

NIH/NINR 

Durability of Adherence in Self-Management of HIV (DASH) 

Major goals: To evaluate the impact of individualized self-management interventions on adherence to highlyactive antiretroviral therapy (HAART) through assessment and modification of an existing intervention intended 

to improve long-term durability of HAART adherence. (Role: Co-Investigator) 

2 R44 AI068543 (Myatt, Chris) 

NIH/NIAID 

09/01/2010-08/31/2012 

Low Cost Multi-Pathogen Laser Diagnostic for HIV and AIDS Co-Infection 

Major goals: Develop a low cost point of care platform for the diagnosis of HIV-1 associated infections in 

resource limited settings. (Role: Co-Investigator) 

1 R24 TW008908-01 (Noormahomed, Emilia) 

NIH 

The Medical Education Partnership Initiative 

09/30/2010-09/29/2015 

Major Goals: Assist Pl in her efforts to develop clinical and translational research capacity at UEM. She will 
Biographical Sketches for each listed Senior/Key Person 3 Page 24

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J

assist Pl in the further development of the translational research laboratory both in terms of equipping the 

laboratory and in terms of providing training for technical staff, trainees and junior faculty in laboratory 

techniques; serve as a member of the UCSD Core Program Faculty and be available to mentor faculty in 

translational research related to HIV transmission, drug resistance and molecular diagnosis. (Role: UCSD Pl of 

Research Component; Co-Investigator) 

R21 AI080353 (Smith, David M.) 

NIH 

04101/2011-03/31/2013 

Pooled Nucleic Acid Testing to Identify HIV Treatment Failure in Mozambique 

To determine the effectiveness of a pooled nucleic acid testing strategy for detecting antiretroviral therapy 

failure and the development of HIV drug resistance in the resource limited setting of Mozambique. (Role: CoInvestigator) 

NIH 2R44 A1070052-04A1 (Schooley) 03/01/2012- 02/28/2013 

Low Cost Laser Diagnostic for CD4 T Cell Counting 

To develop and test in clinical settings a novel, 2-color fluorescence detection method in a low cost point-ofcare platform to identify and quantify CD4+ T cells. 

SSS, Inc Subct (Schooley) 

CRB-DCR01-S-09-00299 IRC004 

11 /01 /2011 - 03/31 /2013 

IRC 004: A Randomized Double-Blind Study Comparing Oseltamivir versus Placebo for the Treatment of 

Influenza in Low Risk Adults 

Completed Research Support 

2 R44AI070052 (Myatt, Chris - Pl) 

NIH/NIAID 

Low Cost Laser Diagnostic for CD4+ T Cell Counting 

09/01 /2009 - 08/31/2011 

Major Goals: To develop a novel, 2-color fluorescence detection method to identify and count CD4+ T-cells. 

(Role: Co-Investigator) 

3 U01 Al27670-18S2 (Benson, Constance - Pl) 

NIH/NIAID 

San Diego AIDS Clinical Trials Unit 

01/01/2000 - 11/30/2006 

Major Goals: The UCSD AIDS Clinical Trials Unit conducts investigator-initiated clinical trials of the NIH/NIAID 

Adult AIDS Clinical Trials Group (AACTG), intended to develop new or improve existing antiviral therapies or 

strategies for the treatment and prevention of HIV infection and its complications in adults and older 

adolescents infected with HIV. (Role: Pl) 

1 R01 AI081321 (Garfein, Richard - Pl) 

NIH/NIDA 

12/01 /2009 - 11/30/2014 

Cross Border Dynamics of MOR/XOR TB Epidemiology by HIV Status in Tijuana, Mexico 

Major Goals: This project aims to describe tuberculosis drug resistance patterns and correlates of drugresistant TB in the cross-border region of northern Mexico and southern California, and to determine the critical 

pathogen transmission dynamics of TB transmission in a high prevalence endemic area in Tijuana and the 

cross-border region of southern California 

Role: Co-Investigator 
Biographical Sketches for each listed Senior/Key Person 4 Page 25

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J

BIOGRAPHICAL SKETCH 

Provide the following information for the key personnel and other significant contributors. 

NAME 

Susan J. Little, M.D. 

eRACOMMONSUSERNAME 

Follow this format for each person. DO NOT EXCEED FOUR PAGES. 

POSITION TITLE 

Professor of Medicine 

EDUCATION/TRAINING (Begin with baccalaureate or other initial professional education, such as nursing, and include postdoctoral training.) 

INSTITUTION AND LOCATION DEGREE YEAR(s) FIELD OF STUDY (if aoolicable) 

Carleton College; Northfield, MN B.A. 1983 Biology 

Washington Univ. School of Med.; St. Louis, MO M.D. 1987 Medicine 

Barnes Hospital; St. Louis, MO Residency 1987-1990 Internal Medicine 

University of California, San Diego; San Diego, CA Fellowship 1990-1993 Infectious Diseases 

A. Personal Statement 

The overarching goal of this application entitled, "Point of Care HIV Antigen/Antibody Diagnostic Device", is to 

a develop a point-of-care HIV-1/2 antibody and p24 antigen detection assay on a single point-of-care platform 

with performance equivalent to FDA-approved laboratory approved 4th generation AHIV-1 Ag/Ab combination 

assays. Specifically, Dr. Little will work with MBio Diagnostics, Inc. to develop and execute the plans to provide 

clinical samples from patients with acute and chronic HIV infection that will include de-identified archived 

samples, as well as newly acquired samples of serum/plasma and potentially other biological fluids from 

patients participating in clinical research at the UCSD Antiviral Research Center (AVRC). Dr. Little will maintain 

her highly successful recruitment program for persons with acute HIV infection (HIV RNA+/Ab-) paired with 

recruitment of appropriate control populations (both HIV+ and HIV-) and provide well-characterized clinical and 

behavioral data linked to regularly collected and banked biological specimens that will be made available to 

MBio and UCSD collaborating investigators. Dr. Little works with a large international collaborative team to 

assess viral replication dynamics and host immune dynamics during acute HIV infection. She directs an NIHfunded Program Project at UCSD to evaluate determinants of HIV transmission. She oversees the integration 

of pathogenesis, translational, and clinical research in the UCSD Acute and Early HIV Infection (AEH) Program 

and mentors post-doctoral research fellows and faculty. Dr. Little is an active investigator in the AIDS Clinical 

Trials Group (ACTG) and has developed numerous ACTG protocols to evaluate new strategies for treatment of 

acute infection. She has been heavily involved in the design of novel approaches to screen and identify acutely 

infected individuals, as well as the evaluation of important questions related to HIV prevention, transmission, 

pathogenesis and treatment. Below, fifteen publications are listed pertinent to the proposed topic, especially #s 

1, 3, 7, 9, 12, 14, 15 as selected from over 80 peer-reviewed manuscripts. 

B. Positions and Honors 

Positions 

10/93-09/97 

10/93-present 

10/97 -06/03 

07 /03-06/05 

0 7 /05-06/09 

07 /09-present 

Associate Physician, University of California, San Diego; San Diego, California 

Attending Physician, Veteran's Administration Medical Center; La Jolla, CA 

Assistant Adjunct Professor of Medicine, University of California, San Diego; San Diego, CA 

Associate Adjunct Professor of Medicine, University of California, San Diego; San Diego, CA 

Associate Professor of Medicine in Residence, University of California, San Diego; San 

Diego, CA 

Professor of Medicine in Residence, University of California, San Diego; San Diego, CA 

Current National/Scientific Committees and Service 

07/01-present World Health Organization HIV Resistance Network (HIVResNet) Scientific Committee 

07/03-present International HIV Drug Resistance Workshop Scientific Committee 

01/04-present Current HIV/AIDS Reports Editorial Board 

10/04-present The HIV Prevention Science Committee: A Joint HPTN-AACTG Committee 

12/04-present Antiviral Therapy Editorial Board 
Biographical Sketches for each listed Senior/Key Person 4 Page 26

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Biographical Sketches for each listed Senior/Key Person 4 Page 27

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J

13. Wertheim JO, Kosakovsky Pond SL, Little SJ, De Gruttola V. Using HIV Transmission Networks to 

Investigate Community Effects in HIV Prevention Trials. PLoS ONE 2011,6:e27775. (PMCID: 

PMC3218056). 

14. Hogan CM, Degruttola V, Sun X, Fiscus SA, Del Rio C, Hare CB, Markowitz M, Connick E, Macatangay B, 

Tashima KT, Kallungal B, Camp R, Morton T, Daar ES, Little S; A5217 Study Team. The setpoint study 

(ACTG A5217): effect of immediate versus deferred antiretroviral therapy on virologic set point in recently 

HIV-1-infected individuals. J Infect Dis. 2012 Jan;205(1):87-96. Epub 2011 Dec15. PubMed PMID: 

22180621; PubMed Central PMCID: PMC3242744. 

15. Karris MY, Anderson CM, Morris SR, Smith OM, Little SJ. The Cost of Missing Acute HIV Infection: 

Testing Antibodies, Antigens, and Nucleic Acids. J Clin Microbial. 2012. Epub 2012/03/24. doi: 

10.1128/JCM.00106-12. Pub Med PMID: 22442319. 

D. Research Support 

Ongoing Research Support 

5 P01 AI074621-05 (Little, Susan J.) 03/15/2008- 02/28/2013 

National Institutes of Health 

Determinants of HIV Transmission 

Major goals: To focus on the identification and systematic evaluation of individuals who have been recently 

infected with HIV and the sexual partners who transmitted HIV to them (Transmission Pairs) to elucidate and 

quantify epidemiologic, behavioral, biologic, virologic, and host factors that contribute to transmission. 

5 R01 AI087164-02 (Woelk, Christopher) 12/1/2009 - 11/30/2014 

National Institutes of Health 

Gene Expression Biomarkers of Immune Recovery in HIV Infected Patients 

Major goals: This research will identify patient genes that contribute to immune recovery and represent new 

targets for drugs that can be used to increase immune recovery. This proposal will also identify which drugs an 

HIV-infected patient should be treated with in order to maximize the extent of their immune recovery. 

3 P01 AI074621-03S1 (UCSD #2010-3954) (Little, Susan J.) 05/03/2010 - 09/26/2012 

National Institutes of Health/NIAID NOT-OD-09-056 (1st Suppl in 03 yr) 

Recovery Act Administrative Supplement 

Determinants of HIV Transmission 

Major goals: To develop and implement the infrastructure to combine data across three highly successful 

acute/early HIV infection cohorts (UCSD, UCSF, and MGH) and utilize the unified dataset to address key 

scientific questions. 

3 P01 AI074621-03S1 (UCSD #2010-3029) (Little, Susan J.) 09/27/2010 - 09/26/2012 

National Institutes of Health/NIAID NOT-00-056 (2nct Suppl in 03 yr) 

Recovery Act Administrative Supplement 

Determinants of HIV Transmission 

Major goals: To estimate and evaluate predictors of universal HIV testing acceptance and use statistical 

techniques applied to HIV sequences to infer a molecular epidemiological transmission network within the 

study population. 

1 DP1 DA034978-01 (Smith, David) 08/01/2012 - 07/31/2017 

National Institutes of Health/NIDA 

Molecular Epidemiology for HIV Prevention for Drug Users and Other Risk Groups 

Major goals: To integrate subject data from local research, clinical and public health entities that are screening 

for and treating HIV infected individuals. 

Completed Research Support 

RN07-SD-702 (Little, Susan J.) 10/01 /2007 - 12/31 /2008 

University of California, California HIV/AIDS Research Program 
Biographical Sketches for each listed Senior/Key Person 4 Page 28

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J

Acute Detection and Early Prevention Trial 

Major goals: To evaluate a program of nucleic acid testing (NAT) to screen for acute HIV infection at Rapid 

HIV testing and counseling sites in the County of San Diego with a plan to coordinate cross-cohort analyses of 

HIV incidence rates, transmission of HIV during primary infection, cost effectiveness of NAT screening, and 

characterization of the demographics of acutely HIV infected clients among participating funded study sites. 

5 U01 AI043638-09 (Richman, Douglas D.) 08/01/1998 - 06/30/2008 

National Institutes of Health 

Southern California Primary Infection Program 

Major goals: A collaborative program between UCSD and Harbor/UCLA Medical Center to combine resources 

and expertise in patient recruitment, clinical trials, and pathogenetic research. This program will identify, 

recruit, and retain subjects with acute or early HIV infection, complete two innovative trials, and answer a 

series of important pathogenesis questions. 

County of San Diego (Smith, David) 11/01/2006 - 02/29/2008 

Early Intervention Services 

Major goals: To develop and implement nucleic acid testing in San Diego County HIV Testing and Counseling 

sites to increase the ability to identify individuals with acute and early HIV infection. 

County of San Diego (Smith, David) 07/01/2007 - 06/30/2008 

Early Intervention, Bridge, and Positive 

Changes Program 

Major goals: These programs serve newly diagnosed HIV infected individuals in San Diego County through a 

multidisciplinary approach, including case management, primary medical and psychological care, health 

education and risk reduction. 

5 R01 NS051132-04 (Little, Susan J.) 07/15/2005 - 06/30/2010 

National Institutes of Health/University of California, San Francisco 

HIV Adaptation in the CNS 

Major goals: A collaborative program between UCSF and UCSD investigators to evaluate of the persistence 

of HIV within functional reservoirs among patients receiving potent antiretroviral therapy. 

3 P01 DA012065-10S1 (Little, Susan J.) 05/01/2009 - 04/30/2011 

National Institutes of Health/NIDA 

NeuroAIDS: Effects of Methamphetamine and HCV 

Major goals: To determine whether methamphetamine use potentiates depletion of gut-associated 

lymphocytes, triggering a cascade of bacterial translocation, and peripheral and central immunopathogenesis 

that leads to early, and potentially persistent, brain injury. 

3 P01 AI074621-02S1 (Little, Susan J.) 09/12/2009- 08/31/2011 

National Institutes of Health 

Recovery Act Diversity Supplement 

Determinants of HIV Transmission 

Major goals: To characterize the risk behaviors and attitudes related to HIV prevention efforts among San 

Diego Latinos and the prevalence of HIV infection within this community, particularly among the non-gay 

identified men who are not historically well represented in HIV surveillance sampling. 
* Last Name

RESEARCH & RELATED BUDGET - SECTION A & B, BUDGET PERIOD 1

* Budget Type:

Enter name of Organization:

* Start Date: * End Date:

* Project Role Base Salary ($)

* Fringe 

Benefits ($) * Funds Requested ($)

9.

8.

7.

6.

5.

4.

3.

2.

1.

Total Funds requested for all Senior Key Persons in the attached file

Total Senior/Key Person

Additional Senior Key Persons:

B. Other Personnel

A. Senior/Key Person

* Number of 

Personnel * Project Role

Acad. 

Months

Sum. 

Months

* Requested 

Salary ($)

Cal. 

Months

Post Doctoral Associates

Graduate Students

Undergraduate Students

Secretarial/Clerical

Total Number Other Personnel Total Other Personnel

Total Salary, Wages and Fringe Benefits (A+B)

Prefix * First Name Middle Name Suffix

* Fringe 

Benefits ($) * Funds Requested ($)

Acad. 

Months

Sum. 

Months

* Requested 

Salary ($)

Cal. 

Months

RESEARCH & RELATED Budget {A-B} (Funds Requested)

* ORGANIZATIONAL DUNS:

OMB Number: 4040-0001 

Expiration Date: 06/30/2011

Project Subaward/Consortium

7

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Dr. Michael J Lochhead PD/PI

Dr. Daniel Nieuwlandt Senior Scientist

1 Kurt Vogel, Engineering Manager 0.60

1 Associate Scientist 12.00

1 Clinical Site Manager 0.60

1 Research Associate 6.00

3 Engineering (Mechanical, Process and Software) 2.40

MBio Diagnostics, Inc.

10/01/2013 09/30/2014 Budget Period 

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Detailed Budget - Year 1 Page 29

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
C. Equipment Description

List items and dollar amount for each item exceeding $5,000

Equipment item * Funds Requested ($)

1.

2.

3.

4.

5.

6.

7.

8.

9.

10.

11. Total funds requested for all equipment listed in the attached file

D. Travel

1. Domestic Travel Costs ( Incl. Canada, Mexico and U.S. Possessions)

2. Foreign Travel Costs

Total Travel Cost

Total Equipment

E. Participant/Trainee Support Costs

1. Tuition/Fees/Health Insurance

2. Stipends

3. Travel

4. Subsistence

5. Other

Number of Participants/Trainees Total Participant/Trainee Support Costs

Funds Requested ($)

Funds Requested ($)

RESEARCH & RELATED Budget {C-E} (Funds Requested)

RESEARCH & RELATED BUDGET - SECTION C, D, & E, BUDGET PERIOD 1

* Budget Type:

Enter name of Organization:

* Start Date: * End Date:

Additional Equipment:

* ORGANIZATIONAL DUNS:

Project Subaward/Consortium

10/01/2013 09/30/2014

MBio Diagnostics, Inc.

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Detailed Budget - Year 1 Page 30

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
F. Other Direct Costs

1. Materials and Supplies

2. Publication Costs

3. Consultant Services

4. ADP/Computer Services

5. Subawards/Consortium/Contractual Costs

6. Equipment or Facility Rental/User Fees

7. Alterations and Renovations

8.

9.

10.

Total Other Direct Costs

G. Direct Costs

Total Direct Costs (A thru F)

H. Indirect Costs Indirect Cost 

Rate (%)

Indirect Cost 

Base ($)

1.

2.

3.

4.

Cognizant Federal Agency

I. Total Direct and Indirect Costs

Total Direct and Indirect Institutional Costs (G + H)

J. Fee

K. * Budget Justification

Indirect Cost Type

Funds Requested ($)

Funds Requested ($)

* Funds Requested ($)

Funds Requested ($)

Funds Requested ($)

RESEARCH & RELATED Budget {F-K} (Funds Requested)

Total Indirect Costs

(Only attach one file.)

(Agency Name, POC Name, and POC Phone Number)

NIH, Division of Financial Services, Frank Mihalovic, 301-443-

1254-Budget Justification.pdf

Other: Tooling and Instrument Components

F&A 45.00

RESEARCH & RELATED BUDGET - SECTION F-K, BUDGET PERIOD 1

* Budget Type:

Enter name of Organization:

* Start Date: * End Date:

* ORGANIZATIONAL DUNS:

Project Subaward/Consortium

10/01/2013 09/30/2014

MBio Diagnostics, Inc.

Budget Period 1

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Detailed Budget - Year 1 Page 31

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
* Last Name

RESEARCH & RELATED BUDGET - SECTION A & B, BUDGET PERIOD 2

* Budget Type:

Enter name of Organization:

* Start Date: * End Date:

* Project Role Base Salary ($)

* Fringe 

Benefits ($) * Funds Requested ($)

9.

8.

7.

6.

5.

4.

3.

2.

1.

Total Funds requested for all Senior Key Persons in the attached file

Total Senior/Key Person

Additional Senior Key Persons:

B. Other Personnel

A. Senior/Key Person

* Number of 

Personnel * Project Role

Acad. 

Months

Sum. 

Months

* Requested 

Salary ($)

Cal. 

Months

Post Doctoral Associates

Graduate Students

Undergraduate Students

Secretarial/Clerical

Total Number Other Personnel Total Other Personnel

Total Salary, Wages and Fringe Benefits (A+B)

Prefix * First Name Middle Name Suffix

* Fringe 

Benefits ($) * Funds Requested ($)

Acad. 

Months

Sum. 

Months

* Requested 

Salary ($)

Cal. 

Months

RESEARCH & RELATED Budget {A-B} (Funds Requested)

* ORGANIZATIONAL DUNS:

OMB Number: 4040-0001 

Expiration Date: 06/30/2011

Project Subaward/Consortium

7

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Dr. Michael J Lochhead PD/PI 0.60

Dr. Daniel Nieuwlandt Senior Scientist 4.80

1 Kurt Vogel, Engineering Manager 0.60

1 Associate Scientist 12.00

1 Clinical Site Manager 3.60

1 Research Associate 6.00

3 Engineering (Mechanical, Process and Software) 2.40

MBio Diagnostics, Inc.

10/01/2014 09/30/2015 Budget Period 

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Detailed Budget - Year 2 Page 32

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
C. Equipment Description

List items and dollar amount for each item exceeding $5,000

Equipment item * Funds Requested ($)

1.

2.

3.

4.

5.

6.

7.

8.

9.

10.

11. Total funds requested for all equipment listed in the attached file

D. Travel

1. Domestic Travel Costs ( Incl. Canada, Mexico and U.S. Possessions)

2. Foreign Travel Costs

Total Travel Cost

Total Equipment

E. Participant/Trainee Support Costs

1. Tuition/Fees/Health Insurance

2. Stipends

3. Travel

4. Subsistence

5. Other

Number of Participants/Trainees Total Participant/Trainee Support Costs

Funds Requested ($)

Funds Requested ($)

RESEARCH & RELATED Budget {C-E} (Funds Requested)

RESEARCH & RELATED BUDGET - SECTION C, D, & E, BUDGET PERIOD 2

* Budget Type:

Enter name of Organization:

* Start Date: * End Date:

Additional Equipment:

* ORGANIZATIONAL DUNS:

Project Subaward/Consortium

10/01/2014 09/30/2015

MBio Diagnostics, Inc.

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Detailed Budget - Year 2 Page 33

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
F. Other Direct Costs

1. Materials and Supplies

2. Publication Costs

3. Consultant Services

4. ADP/Computer Services

5. Subawards/Consortium/Contractual Costs

6. Equipment or Facility Rental/User Fees

7. Alterations and Renovations

8.

9.

10.

Total Other Direct Costs

G. Direct Costs

Total Direct Costs (A thru F)

H. Indirect Costs Indirect Cost 

Rate (%)

Indirect Cost 

Base ($)

1.

2.

3.

4.

Cognizant Federal Agency

I. Total Direct and Indirect Costs

Total Direct and Indirect Institutional Costs (G + H)

J. Fee

K. * Budget Justification

Indirect Cost Type

Funds Requested ($)

Funds Requested ($)

* Funds Requested ($)

Funds Requested ($)

Funds Requested ($)

RESEARCH & RELATED Budget {F-K} (Funds Requested)

Total Indirect Costs

(Only attach one file.)

(Agency Name, POC Name, and POC Phone Number)

NIH, Division of Financial Services, Frank Mihalovic, 301-443-

1254-Budget Justification.pdf

Other: Tooling and Instrument Components

F&A 45.00

RESEARCH & RELATED BUDGET - SECTION F-K, BUDGET PERIOD 2

* Budget Type:

Enter name of Organization:

* Start Date: * End Date:

* ORGANIZATIONAL DUNS:

Project Subaward/Consortium

10/01/2014 09/30/2015

MBio Diagnostics, Inc.

Budget Period 2

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Detailed Budget - Year 2 Page 34

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
BUDGET JUSTIFICATION 

MBio p24 SBIR Phase II Application 

The budget outlined here covers personnel, supplies, and travel, as well as instrument and cartridge 

engineering and testing at MBio Diagnostics. 

Key Personnel 

Michael Lochhead. Dr. Lochhead is Vice President at MBio Diagnostics serves as PI on the project. Dr. 

Lochhead has been managing the immunoassay product development at the company since July 2007. He 

provides overall program management and strategy and he will commit 5% effort to this project. 

Daniel Nieuwlandt. Dr. Nieuwlandt is a Senior Scientist at MBio and he will lead assay development efforts 

under Aims 1 – 3. Dr. Nieuwlandt is an expert in bioconjugation techniques and assay development, and he 

was the senior scientist on Phase I of this program. He commits 40% effort to the project (4.8 calendar 

months). 

Other Personnel 

Associate Scientist. MBio will assign one of its Associate Scientists to work full time on the project 

through the program duration. Candidates include Katie Todorof, Phil Papst and Greg Husar, all of whom 

are experience immunoassay development scientists. 

Research Associate. MBio will assign a Research Associate at 50% effort throughout the project. The 

role of the Research Associate will provide basic laboratory support, including running assays, preparing 

reagents, etc. 

Kurt Vogel. Dr. Vogel is Director of Engineering at MBio Diagnostics. His group is responsible for the 

instrument hardware and software development as well as disposable cartridge manufacturing. Dr. Vogel 

will coordinate system modifications associated with the p24 assay. He will commit 5% effort to this project. 

Kevin Moll. Dr. Moll is Senior Scientist at MBio. In addition to providing optical design expertise, he serves 

as MBio’s primary in-house software developer. Dr. Moll develops the company’s proprietary image analysis 

algorithms and coordinates system software development. He will commit 33% effort to this project. 

Keagan Rowley. Mr. Rowley is the lead mechanical design engineer on the project. He is responsible for 

modifying MBio cartridge designs in support of the p24 project. He will also be responsible for part design, 

vendor management, and builds for Aim 2. Mr. Rowley has a 20% percent commitment to the project. 

Process Engineer. The Process Engineer is a new hire at MBio, scheduled for September 2012. The 

process engineer will perform process development research related to cartridge integration tasks under 

Aim 2. The process engineer will dedicate 30% effort throughout the project. 

Clinical Site Manager. During year 2 when clnical evaluation is taking place at UCSD, MBio will support a 

clinical site manager who works on data management, logistics, sample transfers, etc. with the UCSD 

clinical team. This position will be at a 5% level during year 1, increasing to 30% during year 2. 

Equipment 

No capital equipment is requested under this award. 

Budget Justification Page 35

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
USupplies 

The Supplies budget worksheets are provided in the tables below. 

Antibody and Reagent Cost Calculation: 

Reference test kits are a substantial part of the budget. For example, 4th generation HIV Ag/Ab Combo 

assays (e.g., Bio-Rad) cost > $ per microplate. Reference test costing is based on experience from 

Phase I. 

Other: Tooling and Instrument Components 

In addition to the standard laboratory and cartridge supplies listed above, we designate tooling and supplies 

for instrument development in their own “other” category. 

The Cartridge Mold line item assumes that during each year we will do one revision of the molded cartridge. 

MBio works with multiple molding vendors. The current SnaEsi cartridge body mold was $ (steel tool 

to hold polish and throughput), and we use that number as an estimate for revisions. The cartridge also has 

a lid which will be based on less expensive aluminum tooling, such as through Protomold. We budget 

$ for the lid mold and two iterations at Protomold. Total mold budget with run time costs is estimated 

at $ during year 1. 

We will work with custom blister pack vendors to incorporate on board liquid reagents for the final wash step 

outlined under Aim 2. MBio has experience with several of these vendors. 

Instrument builds for the program will be at $ per unit, with one built in year 1 and two in year 2. 

UTravel 

The travel budget anticipates two trips per year. One will be for two people to visit the clinical collaborators 

at UCSD and coordinate the field evaluation. For each person, we budget $ transportation (air and 

ground), $ lodging, and $ meals ($ per person). The total for two people is $ 

Consultants 

We request funding for specific regulatory support associated with the 4th Gen HIV product. Because such 

a test will require a full pre-market approval (PMA) through CBER at FDA, we will be looking for specific 

guidance from our regulatory consultants. MBio currently engages Myraqa for strategic regulatory 

consulting. We budget $ per year for these consulting services. 

Fee

We include the allowed 7% fee in our budget. The 7% is calculated on the MBio Direct and Indirect costs, 

excluding the UCSD subaward. 

Supplies Item Subtotal Item Total

Antigens and Antibodies $ See calculation table

Labw are (gloves, tubes pipets, etc.) $ $ /month based on past experience

Cartridges (materials & labor) $ $ Cost includes materials and labor

Reference Test Kits $ 

Supplies: $ 

ANTIGEN/ANTIBODY CALCULATOR:

Analytes/ Reagents: 6 antibodies

Labeling kits: 2 e.g., Invitrogen

Repeat orders: 3

Price per order: $ average

TOTAL: $

Other: Tooling and Instrument Components

Cartridge Molds $ 

Blister pack vendor support $ 

SnapEsi System (dedicated to p24) $ 

Other: $ 

Budget Justification Page 36

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Section A, Senior/Key Person

Section C, Equipment

RESEARCH & RELATED BUDGET - Cumulative Budget

Section D, Travel

Domestic

Section E, Participant/Trainee Support Costs

Foreign

Tuition/Fees/Health Insurance

Stipends

Travel

Subsistence

Other

Number of Participants/Trainees

1.

2.

3.

4.

5.

6.

1.

2.

Section F, Other Direct Costs

1. Materials and Supplies

2. Publication Costs

3. Consultant Services

4. ADP/Computer Services

5. Subawards/Consortium/Contractual Costs

6. Equipment or Facility Rental/User Fees

7. Alterations and Renovations

8.

9.

10.

Totals ($)

Total Number Other Personnel

Total Salary, Wages and Fringe Benefits (A+B)

Other 1

Other 2

Other 3

Section B, Other Personnel

Section J, Fee

Section I, Total Direct and Indirect Costs (G + H)

Section H, Indirect Costs

Section G, Direct Costs (A thru F)

14

	


 		

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Cumulative Budget Page 37

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED BUDGET - SECTION A & B, BUDGET PERIOD 1

* ORGANIZATIONAL DUNS:

* Budget Type: ❍ Project ● Subaward/Consortium

Enter name of Organization: The Regents of the Univ. of Calif., San Diego

* Start Date: 10-01-2013 * End Date: 09-30-2014 Budget Period: 1

A. Senior/Key Person

Prefix * First Name Middle Name * Last Name Suffix * Project Role Base Salary

($)

Cal.

Months

Acad.

Months

Sum.

Months

* Requested

Salary ($)

* Fringe

Benefits ($)

* Funds Requested ($)

1. Constance Benson MD Consortium PI 0.60

2. Susan Little MD Co-Investigator 0.30

Total Funds Requested for all Senior Key Persons in the attached file

Additional Senior Key Persons: File Name: Mime Type: Total Senior/Key Person 18,409.00

B. Other Personnel

* Number of

Personnel

* Project Role Cal.

Months

Acad.

Months

Sum.

Months

* Requested

Salary ($)

* Fringe

Benefits

* Funds Requested

($)

Post Doctoral Associates

Graduate Students

Undergraduate Students

Secretarial/Clerical

1 Jill Kunkel, Research Nurse 1.20

1 Denise Lovec-Jenkins, Regulatory Specialist 1.20

2 Total Number Other Personnel Total Other Personnel

Total Salary, Wages and Fringe Benefits (A+B)

RESEARCH & RELATED Budget {A-B} (Funds Requested)

Tracking Number: GRANT11392590 OMB Number: 4040-0001

Expiration Date: 06/30/2011

Subaward 1 Page 38

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED BUDGET - SECTION C, D, & E, BUDGET PERIOD 1

* ORGANIZATIONAL DUNS:

* Budget Type: ❍ Project ● Subaward/Consortium

Enter name of Organization: The Regents of the Univ. of Calif., San Diego

* Start Date: 10-01-2013 * End Date: 09-30-2014 Budget Period: 1

C. Equipment Description

List items and dollar amount for each item exceeding $5,000

Equipment Item * Funds Requested ($)

Total funds requested for all equipment listed in the attached file

Total Equipment

Additional Equipment: File Name: Mime Type:

D. Travel Funds Requested ($)

1. Domestic Travel Costs ( Incl. Canada, Mexico, and U.S. Possessions)

2. Foreign Travel Costs

Total Travel Cost

E. Participant/Trainee Support Costs Funds Requested ($)

1. Tuition/Fees/Health Insurance

2. Stipends

3. Travel

4. Subsistence

5. Other:

Number of Participants/Trainees Total Participant/Trainee Support Costs

RESEARCH & RELATED Budget {C-E} (Funds Requested)

Tracking Number: GRANT11392590 OMB Number: 4040-0001

Expiration Date: 06/30/2011

Subaward 1 Page 39

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED BUDGET - SECTIONS F-K, BUDGET PERIOD 1

* ORGANIZATIONAL DUNS:

* Budget Type: ❍ Project ● Subaward/Consortium

Enter name of Organization: The Regents of the Univ. of Calif., San Diego

* Start Date: 10-01-2013 * End Date: 09-30-2014 Budget Period: 1

F. Other Direct Costs Funds Requested ($)

1. Materials and Supplies

2. Publication Costs

3. Consultant Services

4. ADP/Computer Services

5. Subawards/Consortium/Contractual Costs

6. Equipment or Facility Rental/User Fees

7. Alterations and Renovations

8. Communications and Computing

9. Outpatient Care Costs

Total Other Direct Costs

G. Direct Costs Funds Requested ($)

Total Direct Costs (A thru F)

H. Indirect Costs

Indirect Cost Type Indirect Cost Rate (%) Indirect Cost Base ($) * Funds Requested ($)

1. MTDC, On Campus Rate 55.00

Total Indirect Costs

Cognizant Federal Agency

(Agency Name, POC Name, and POC Phone Number)

DHHS Region IX 415-437-7820

I. Total Direct and Indirect Costs Funds Requested ($)

Total Direct and Indirect Institutional Costs (G + H)

J. Fee Funds Requested ($)

K. * Budget Justification File Name: 1234-UCSDBudgetJustification.pdf Mime Type: application/pdf

(Only attach one file.)

RESEARCH & RELATED Budget {F-K} (Funds Requested)

Tracking Number: GRANT11392590 OMB Number: 4040-0001

Expiration Date: 06/30/2011

Subaward 1 Page 40

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED BUDGET - SECTION A & B, BUDGET PERIOD 2

* ORGANIZATIONAL DUNS: 8043557900000

* Budget Type: ❍ Project ● Subaward/Consortium

Enter name of Organization: The Regents of the Univ. of Calif., San Diego

* Start Date: 10-01-2014 * End Date: 09-30-2015 Budget Period: 2

A. Senior/Key Person

Prefix * First Name Middle Name * Last Name Suffix * Project Role Base Salary

($)

Cal.

Months

Acad.

Months

Sum.

Months

* Requested

Salary ($)

* Fringe

Benefits ($)

* Funds Requested ($)

1. Constance Benson MD Consortium PI 0.60

2. Susan Little MD Co-Investigator 0.30

Total Funds Requested for all Senior Key Persons in the attached file

Additional Senior Key Persons: File Name: Mime Type: Total Senior/Key Person 18,638.00

B. Other Personnel

* Number of

Personnel

* Project Role Cal.

Months

Acad.

Months

Sum.

Months

* Requested

Salary ($)

* Fringe

Benefits

* Funds Requested

($)

Post Doctoral Associates

Graduate Students

Undergraduate Students

Secretarial/Clerical

1 Jill Kunkel, Research Nurse 1.20

1 Denise Lovec-Jenkins, Regulatory Specialist 1.20

2 Total Number Other Personnel Total Other Personnel

Total Salary, Wages and Fringe Benefits (A+B)

RESEARCH & RELATED Budget {A-B} (Funds Requested)

Tracking Number: GRANT11392590 OMB Number: 4040-0001

Expiration Date: 06/30/2011

Subaward 1 Page 41

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED BUDGET - SECTION C, D, & E, BUDGET PERIOD 2

* ORGANIZATIONAL DUNS:

* Budget Type: ❍ Project ● Subaward/Consortium

Enter name of Organization: The Regents of the Univ. of Calif., San Diego

* Start Date: 10-01-2014 * End Date: 09-30-2015 Budget Period: 2

C. Equipment Description

List items and dollar amount for each item exceeding $5,000

Equipment Item * Funds Requested ($)

Total funds requested for all equipment listed in the attached file

Total Equipment

Additional Equipment: File Name: Mime Type:

D. Travel Funds Requested ($)

1. Domestic Travel Costs ( Incl. Canada, Mexico, and U.S. Possessions)

2. Foreign Travel Costs

Total Travel Cost

E. Participant/Trainee Support Costs Funds Requested ($)

1. Tuition/Fees/Health Insurance

2. Stipends

3. Travel

4. Subsistence

5. Other:

Number of Participants/Trainees Total Participant/Trainee Support Costs

RESEARCH & RELATED Budget {C-E} (Funds Requested)

Tracking Number: GRANT11392590 OMB Number: 4040-0001

Expiration Date: 06/30/2011

Subaward 1 Page 42

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED BUDGET - SECTIONS F-K, BUDGET PERIOD 2

* ORGANIZATIONAL DUNS:

* Budget Type: ❍ Project ● Subaward/Consortium

Enter name of Organization: The Regents of the Univ. of Calif., San Diego

* Start Date: 10-01-2014 * End Date: 09-30-2015 Budget Period: 2

F. Other Direct Costs Funds Requested ($)

1. Materials and Supplies

2. Publication Costs

3. Consultant Services

4. ADP/Computer Services

5. Subawards/Consortium/Contractual Costs

6. Equipment or Facility Rental/User Fees

7. Alterations and Renovations

8. Communications and Computing

9. Outpatient Care Costs

Total Other Direct Costs

G. Direct Costs Funds Requested ($)

Total Direct Costs (A thru F)

H. Indirect Costs

Indirect Cost Type Indirect Cost Rate (%) Indirect Cost Base ($) * Funds Requested ($)

1. MTDC, On Campus Rate 55.00

Total Indirect Costs

Cognizant Federal Agency

(Agency Name, POC Name, and POC Phone Number)

DHHS Region IX 415-437-7820

I. Total Direct and Indirect Costs Funds Requested ($)

Total Direct and Indirect Institutional Costs (G + H)

J. Fee Funds Requested ($)

K. * Budget Justification File Name: 1234-UCSDBudgetJustification.pdf Mime Type: application/pdf

(Only attach one file.)

RESEARCH & RELATED Budget {F-K} (Funds Requested)

Tracking Number: GRANT11392590 OMB Number: 4040-0001

Expiration Date: 06/30/2011

Subaward 1 Page 43

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
RESEARCH & RELATED BUDGET - Cumulative Budget

Totals ($)

Section A, Senior/Key Person

Section B, Other Personnel

Total Number Other Personnel 4

Total Salary, Wages and Fringe Benefits (A+B)

Section C, Equipment

Section D, Travel

1. Domestic

2. Foreign

Section E, Participant/Trainee Support Costs

1. Tuition/Fees/Health Insurance

2. Stipends

3. Travel

4. Subsistence

5. Other

6. Number of Participants/Trainees

Section F, Other Direct Costs

1. Materials and Supplies

2. Publication Costs

3. Consultant Services

4. ADP/Computer Services

5. Subawards/Consortium/Contractual Costs

6. Equipment or Facility Rental/User Fees

7. Alterations and Renovations

8. Other 1

9. Other 2

10. Other 3

Section G, Direct Costs (A thru F)

Section H, Indirect Costs

Section I, Total Direct and Indirect Costs (G + H)

Section J, Fee

Tracking Number: GRANT11392590 OMB Number: 4040-0001

Expiration Date: 06/30/2011

Subaward 1 Page 44

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
UCSD Budget Justification

PERSONNEL:

Constance Benson, M.D. (Consortium PI – 0.60 Calendar Months). Dr. Benson is a 

Professor of Medicine at UCSD and Director of the UCSD Antiviral Research Center. She will 

coordinate and supervise all research activities for the project, and will be responsible for 

recruitment and enrollment of research participants. She will recruit individuals from participants 

in clinical and translational research studies at the AVRC, obtain informed consent and abstract 

correlative clinical information for the SBIR project. 

Susan Little, M.D. (Consortium Co-Investigator – 0.3 Calendar Months). Dr. Little is a 

Professor of Medicine at UCSD and a co-investigator of the UCSD Antiviral Research Center. 

She directs the Acute and Early HIV Infection (AEH) Program at the AVRC, and coordinates the 

clinical and specimen repository for the AEH Program. She will coordinate access to stored 

samples for assay development, and will assist Dr. Benson in participant recruitment, obtaining 

informed consent, and abstracting correlative clinical information for the SBIR project.

Jill Kunkel, R.N. (Study Coordinator and Chief Research Nurse of the Antiviral Research 

Center – 1.20 Calendar Months). Ms. Kunkel will assist Dr. Benson in participant recruitment 

as well as in logistics of blood acquisition and data extraction. 

Denise Lovec-Jenkins (Regulatory Specialist – 1.20 Calendar Months). Ms. Lovec-Jenkins 

will coordinate regulatory submissions and renewals. 

OTHER DIRECT COSTS: 

Supplies ($ are budgeted to cover costs of laboratory disposables such as gloves, blood 

collection materials, etc.

Patient care costs ($ are budgeted to cover costs of reference testing using HIV EIA 

and NAT testing for 200 samples prospectively collected for the project (HIV EIA x 200 samples 

x $ per sample = $ HIV NAT x 200 samples x $ per sample = $

NGN/Communications/Computing ($ costs have been included for telephone and 

associated voice and data communication charges which are directly related to the individuals 

working on the project.

Subaward 1 Budget Justification Attachment Page 45

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
SBIR/STTR Information OMB Number: 4040-0001 

Expiration date: 06/30/2011 

* Program Type (select only one)

* SBIR/STTR Type (select only one)

* 1a. Do you certify that at the time of award your organization will meet the eligibility criteria for a small business as defined in the funding 

opportunity announcement?

* 2. Does this application include subcontracts with Federal laboratories or any other Federal Government agencies?

* If yes, insert the names of the Federal laboratories/agencies:

* 3. Are you located in a HUBZone? To find out if your business is in a HUBZone, use the mapping utility provided by the Small Business 

* 4. Will all research and development on the project be performed in its entirety in the United States?

* Explanation:

* 5. Has the applicant and/or Program Director/Principal Investigator submitted proposals for essentially equivalent work under other 

Federal program solicitations or received other Federal awards for essentially equivalent work? 

* If yes, insert the names of the other Federal agencies:

* 6. Disclosure Permission Statement: If this application does not result in an award, is the Government permitted to disclose the title of 

your proposed project, and the name, address, telephone number and e-mail address of the official signing for the applicant organization, 

to organizations that may be interested in contacting you for further information (e.g., possible collaborations, investment)?

* 7. Commercialization Plan: If you are submitting a Phase II or Phase I/Phase II Fast-Track Application, include a 

Commercialization Plan in accordance with the agency announcement and/or agency-specific instructions.

* Attach File:

Questions 1-7 must be completed by all SBIR and STTR Applicants:

If no, provide an explanation in an attached file.

Administration at its web site: http://www.sba.gov 

SBIR STTR

Both (See agency-specific instructions to determine whether a particular agency allows a single submission for both SBIR and STTR)

Phase I Phase II

Fast-Track (See agency-specific instructions to determine whether a particular agency participates in Fast-Track)

Yes

No

30

Yes

No

Yes

No

Yes

No

Yes

No

Yes

No

Add Attachment Delete Attachment View Attachment

1252-Commercialization Plan p24 Add Attachment Delete Attachment View Attachment

* 1b. Anticipated Number of personnel to be employed at your organization at the time of award.

	


 		

 !"  #

SBIR/STTR Information Page 46

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
* 8. Have you received SBIR Phase II awards from the Federal Government? If yes, provide a company commercialization history in 

accordance with agency-specific instructions using this attachment. 

* Attach File:

* 9. Will the Project Director/Principal Investigator have his/her primary employment with the small business at the time of award?

* 10. Please indicate whether the answer to BOTH of the following questions is TRUE:

(1) Does the Project Director/Principal Investigator have a formal appointment or commitment either with the small business directly 

(as an employee or a contractor) OR as an employee of the Research Institution, which in turn has made a commitment to the 

small business through the STTR application process; AND 

(2) Will the Project Director/Principal Investigator devote at least 10% effort to the proposed project?

* 11. In the joint research and development proposed in this project, does the small business perform at least 40% of the work and the research 

institution named in the application perform at least 30% of the work?

SBIR-Specific Questions:

STTR-Specific Questions:

SBIR/STTR Information

Questions 8 and 9 apply only to SBIR applications. If you are submitting ONLY an STTR application, leave questions 8 and 9 blank and proceed to 

question 10.

Questions 10 and 11 apply only to STTR applications. If you are submitting ONLY an SBIR application, leave questions 10 and 11 blank.

1253-Commercial History - Less Add Attachment Delete Attachment View Attachment

Yes

No

Yes

No

Yes

No

Yes

No

	


 		

 !"  #

SBIR/STTR Information Page 47

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
COMMERCIALIZATION PLAN 

Executive Summary 

The mission of MBio Diagnostics, Inc. is to improve health outcomes by providing the right 

information, in the right place, at the right time. Our initial focus is on low cost point-of-care 

technology emphasizing simplicity while providing laboratory quality results.

The company is advancing two product applications based on its proprietary fluorescence assay 

platform. Both have initial products in the in the field of HIV/AIDS. The first is a multiplexed 

serodiagnostic tool that either individually or simultaneously provides diagnostic results for HIV 

and common AIDS opportunistic infections (hepatitis B & C, syphilis, HHV-8, etc.), all from a 

single drop of blood. The technology for p24 viral antigen detection in this proposal will enable 

a high sensitivity HIV test that can detect the presence of virus soon after the onset of viremia. 

The technology will be included in a antenatal screening panel in development at MBio, a panel 

that will screen for HIV, HBV, syphilis, and anemia all on a single device with a single 10ul 

sample. The second product is a simplified cytometer for routine enumeration of CD4 cells for 

management of HIV-infected individuals. HIV-1 mediated CD4 cell destruction is the central 

immunologic feature of HIV-1 infection. Thus, the CD4 count is a critical measurement in initial 

disease staging, in monitoring antiretroviral therapy and in managing primary and secondary 

prophylaxis for opportunistic infections. 

This Phase 2 application seeks funding for an innovative approach to serological testing for HIV 

that shows promise in detecting HIV at the earliest stages, yet with the high specificity that 

makes it a valued test for screening. The company is seeking SBIR Phase 2 funding to develop 

the technique and demonstrate its utility in the clinic. At the same time, MBio is pursuing 

strategic partnerships and venture capital investment that will be used to drive development 

through manufacturing, regulatory filings, sales, and distribution. 

MBio scientists and engineers are driven to make an impact on health, and a simple rapid tool 

that allows detection of HIV at the earliest, most infectious stages can greatly slow the spread of 

the disease. In general, the sensitive and specific detection of circulating proteins has value in 

many diagnostic applications, in both infectious disease (e.g. HBV surface antigen, cryptococcal 

antigen) and non-infectious disease (e.g. cardiac troponin). 

A. Value of the SBIR/STTR Project, Expected Outcomes, and Impact 

Value 

The research that will be conducted under this grant will lead to a low-cost, ultra-sensitive POC 

HIV test, enabling detection of disease at its earliest stages in settings where HIV prevalence is 

highest. In general, the core technology serves as a platform for the development of other 

critical diagnostics tools, such as diagnostic panels for hepatitis, other infectious diseases, and 

non-communicable diseases. The value of all the applications in aggregate is substantial; here 

we will focus on the value of a highly sensitive POC HIV test that can directly detect acute 

infections in the clinic, and describe the overall market for products that could be enabled by the 

technology. 

In the United States, it is estimated that there are 1.3 million individuals infected with HIV, and 

another 50,000 per year are newly infected. Approximately 250,000 individuals do not know 

their status, and those that do not know drive about half of the new infections, according to CDC 

estimates. The financial cost of treating HIV over the course of a lifetime is estimated at 

Commercialization Plan Page 48

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
$ representing $ billion dollars for the current caseload, and increasing by $ 

billion each year. This is just in the US, and similarly staggering sums can be calculated for the 

caseload worldwide of nearly 33 million individuals. In short, the value of a targeted screening 

program within the most impacted communities in the US, and leading to a decrease of 10,000 

infections per year represents a cost savings of $6 billion. The CDC has advocated both wide 

screening approaches, such as for any person seeking care at an emergency department, and 

focused screening for populations at highest risk. The goal is to directly impact the new 

infection rate by increasing awareness of HIV status. A technology that is highly sensitive to 

acute infections (~30x increase in infectivity during the acute phase of the disease), can enable 

immediate contact tracing (as opposed to a weeks-long delay for getting lab-based nucleic acid 

testing results), and is available in a format that is simple to run and that allows many individuals 

to be screened in parallel with a low-cost device (as opposed to any of the POC nucleic acid 

devices under development), is a clear enabling technology for such a screening program. 

Expected Outcomes 

The direct outcome of this program is a highly sensitive, specific, and affordable technology for 

detecting and quantifying proteins in a biological sample in a clinic or other point-of-care setting. 

This technology enables multiple products, including a 4th generation HIV test, a p24 antigen 

quantification test for ART therapy monitoring, a cardiac troponin test, an improved hepatitis B 

screening test, and a combination test for antenatal screening including HIV, hepatitis B, and 

syphilis, all on a single device. Each of these tests can make a significant impact on health, 

both in the US and in a global context. 

Impact 

To understand the impact of this proposal, it is necessary to put the resulting products in the 

context of the overall platform under development at MBio. While each test we develop has 

intrinsic value, there is an even greater value for an overall platform that delivers a menu of 

tests at the point of care. In the case of HIV, the MBio platform can simultaneously screen for 

multiple diseases including HIV, then provide a confirmation test that separately reports each of 

several markers for HIV, then perform a CD4 test, screen for multiple co-infections, and 

measure viral load for a confirmed case of HIV. Each of these indications is under development 

at MBio, and shows how a single platform can provide a comprehensive set of diagnostic 

information. A successful completion of this development, followed by commercialization and 

market acceptance, would lead to critical enabling of HIV ART therapy throughout the world. It 

would permit wider distribution of such therapy, helping to limit the tragedy wrought by HIV. In 

the US, an improved HIV screening technology that is sensitive during the viremic stage of the 

disease can have a real impact on the ongoing epidemic and its human and financial costs. 

In the broader context of society, there is a critical need to drive efficiency in the healthcare 

system—and if this is not achieved, it will bankrupt the United States. Increased efficiency will 

demand earlier and more accurate interventions—the cornerstones of personalized medicine. 

B. Company 

MBio Diagnostics, Inc. was established in 2007 as a division of Precision Photonics Corporation 

(PPC), a leading manufacturer of laser technology, located in Boulder, Colorado. MBio was 

spun off as an independent company in 2010. Founded by optical measurement scientists from 

Nobel-prize winning laboratories at the University of Colorado, MBio’s mission is to improve 

health outcomes by providing the right information, in the right place, at the right time. To 

complement the instrumentation expertise of the company’s founders, MBio has assembled a 

team of diagnostic industry veterans and has established strong relationships with a world-class 

Commercialization Plan Page 49

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
group of clinical collaborators. The company also has an active Silicon Valley based investor 

group that has provided seed financing and ongoing business guidance. MBio Diagnostics, Inc. 

is now a 24 person, privately held company based in Boulder. 

Leadership 

The company’s leadership team is comprised of:

Dr. Christopher Myatt, PhD – CEO – Chris has over 20 years of experience in technology 

development and building companies that capitalize on advanced measurement technology and

scalable manufacturing. He founded Precision Photonics Corporation, an optical device design 

and manufacturing business that was successfully sold in 2012. The MBio technology was 

initially developed at Precision Photonics, and then spun out into a separate company prior to 

the sale of the parent company. Chris earned his PhD at University of Colorado/JILA, where his 

research work under Prof. Carl Wieman culminated in the first creation of Bose-Einstein 

Condensation (BEC) in Prof. Wieman’s laboratory. Prof. Wieman was awarded the 2001 Nobel 

Prize in Physics for BEC. 

Dr. Michael Lochhead – Vice President – Mike has over 10 years experience leading 

development teams in the life sciences and diagnostics field, with successful products on the 

market. He has led the growth of MBio from a two-person research project within Precision 

Photonics to a 24-person independent diagnostics company. He brings critical expertise in both 

the science and project management of a multi-disciplinary team. Dr. Lochhead completed his 

post-doctoral training in the Bioengineering Department at the University of Washington, and 

has a PhD in Chemical Engineering from the University of the Wisconsin, Madison. 

Michael Seely – Chief Commercial Officer – Michael is an expert at building international 

commercial organizations, having founded and run medical technology businesses in South 

America, Southeast Asia, and Europe. Michael will lead our efforts to commercialize MBio 

products world-wide.

Cindi Moore – Controller – Cindi has over 20 years experience in accounting and finance, 

including biotech firms and manufacturing companies. 

In addition to internal company resources, MBio has partnered with Dr. Robert Schooley and Dr. 

Constance Benson from University of California, San Diego (UCSD) Medical School. Drs. 

Schooley and Benson provide clinical guidance to the company and will be working with MBio 

on validation of the SnapCountTM technology. Dr. Schooley is Head of the Division of Infectious 

Diseases at UCSD and currently serves on the Scientific Advisory Board of the U.S. President’s 

Emergency Plan for AIDS Relief (PEPFAR). Through his affiliation with the NIH AIDS Clinical 

Trials Group (ACTG), his international network includes clinical research sites in Latin America 

(Peru and Brazil), the Caribbean (Haiti), Africa (Mozambique, South Africa, Botswana, 

Zimbabwe, Malawi, Zambia, Kenya and Uganda), and Asia (India and Thailand). Working with 

investigators from these sites he has developed an initial scientific agenda for the ACTG 

International Program. One of the major goals of this agenda is the development of accurate 

and inexpensive point-of-care diagnostics that can be deployed in resource limited settings 

where disease management is often syndromic because of the lack of local diagnostic 

capabilities. In addition, he collaborates with investigators in Kenya (Dr. Fred Sawe, Director, 

Kericho Walter Reed Program), Zimbabwe (Dr. James Hakim, Chairman, Department of 

Medicine, University of Zimbabwe) and Brazil (Dr. Roberto Badaro, Chief, Division of Infectious 

Diseases, Federal University of Bahia). 

Commercialization Plan Page 50

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Funding History

MBio’s funding to date is from four sources: research grants and contracts, funding by former 

parent company Precision Photonics Corporation (PPC), private investment, and product sales. 

To date, the company has received research grants and development contracts totaling over 

 million, with million remaining on various contracts and grants. This research funding 

not only provides key development resources, but winning competitive proposals is an indirect 

validation of MBio’s technology and approach.

From the time the group was founded through the end of 2010, parent company PPC provided 

significant direct and in-kind support to the MBio effort, covering major costs not addressed by 

research contracts and grants, including intellectual property investment as well as facility 

expansion, business development, and overhead. In addition, MBio has a core group of private 

investors, led by Milton Chang of Incubic, who have recently invested into MBio to 

achieve initial regulatory approvals and product launch. This financing was a re-investment of a 

portion of the proceeds from the recent sale of PPC, representing a significant vote of 

confidence in the MBio team. MBio will continue to seek investment financing to expand its 

product offerings and bring those products to market. 

Finally, MBio has begun shipping products, currently for research use only, including a panel for 

detecting shellfish toxins in a food safety application. First regulatory approvals for diagnostic 

tests are expected in with growing product sales beginning in the timeframe. 

This growing business will continue to fund product improvements and corporate development. 

Business Strategy 

MBio has a platform diagnostic technology that offers multiplexing, quantitation, high sensitivity, 

and data connectivity in a simple, affordable package that enables point-of-care testing. As a 

platform technology, there are many applications that can be developed. As a small company, 

we have limited resources for addressing this multitude of applications. Thus our business 

strategy is to complete the technology development, demonstrate its utility in certain 

applications that the company will develop and bring to market, and look to partners to develop 

other applications, either as licensees or acquirers of the technology for specific applications. 

MBio has chosen to focus our internal development on unmet market needs within infectious 

disease testing. MBio is developing initial applications in testing for HIV, hepatitis, influenza, 

and syphilis with external R&D funding and has established strong and active working 

relationships with key opinion leaders in the global health field. The technology proposed here 

will significantly advance our product offering by enabling market leading performance in HIV, 

and open up other high sensitivity applications in fields such as cardiology and rheumatology. 

MBio is in active collaboration or discussion for development of a variety of other applications of 

our technology. Examples include an ongoing development of a food safety application, a joint 

project to develop a TB serology diagnostic, and several discussions about development of 

specific panels as companion diagnostics for therapeutic. This last application is particularly 

exciting, as it would enable personalized medicine directly in the clinic, and speed up the clinical 

decision process. 

Regulatory Strategy 

In anticipation of United States FDA and European (CE Mark) regulatory filings, MBio is 

implementing Quality Systems under 21 CFR 820 and ISO 13485. MBio’s initial products are 

being developed under Design Control, and we anticipate that our new pilot production facility 

Commercialization Plan Page 51

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
will be cGMP compliant in MBio’s regulatory strategy is to gain approval in Europe and 

other markets outside of the US first, and then bring the technology for approval in the US. Our 

first indication, a CD4 enumeration assay, is a relatively low risk device that can be self-certified 

in Europe (expected and faces a 510(k) process in the US (expected The HIV 4th

generation test anticipated by the present proposal would require more elaborate regulatory 

approvals, since an HIV diagnostic test is included in Annex 2a under the European system, 

and requires a PMA for approval in the US. These regulatory hurdles will be addressed starting 

in 

Reimbursement Strategy 

MBio has chosen to develop first products for indications with established CPT codes and 

purchasing channels. This focus on known reimbursement and purchasing reduces risk in the 

business plan. The products anticipated from the proposed effort, such as a 4th generation 

rapid HIV test, or adjacent products such as a HBV diagnostic or a cardiac troponin test, all 

have well defined reimbursement codes. For the HIV test, there is a chance for code stacking, 

since the test reports both the antibody and antigen status. 

C. Market, Customer, and Competition 

Market 

MBio estimates that the global market for in-vitro diagnostics exceeds $75B annually. MBio 

intends to establish itself as a leader in the professional point-of-care testing (POCT) segment 

of this market, which is estimated to be, over all applications, between $4B and $5B. The initial 

markets for the MBio technology are defined by the need for low-cost rapid tests for HIV, both 

diagnosing the disease itself and its common co-infections, and tests to manage therapy. The 

HIV/AIDS testing market, both POC and lab-based, is estimated to be greater than US$1B 

annually and growing at a rate of 15% per year. This growth is due to the increased prevalence 

of the disease and the increased funding being applied to testing patients worldwide, while 

being moderated by price decreases as more competitors enter the space. 

Analysts at the Clinton Health Access Initiative (CHAI) are developing an analysis of the global 

demand for point-of-care (POC) testing. CHAI has shared preliminary results of this analysis 

with MBio. We have supplemented this analysis with several market reports, and with internal 

analysis. We are projecting the demands to 2017, given the long lead times for regulatory 

approval and market acceptance. Factored in are the initiatives that are underway to increase 

HIV screening (e.g. US CDC mandate to screen all emergency department visitors) and HIV 

therapy (e.g. the ―15x15 Initiative‖, to put 15 million persons on HIV ART by 2015). The 

standard of care for HIV ART monitoring in the US is a viral load and CD4 measurement 4 times 

per year, while currently in the developing world 1-2 CD4 tests per year are standard. However, 

the desire is to increase this monitoring, to 2x per year for viral monitoring (either a viral load 

test or a semi-quantitative ―viral monitor‖ test that indicates viral replication when a signal is 

above a set threshold), and 1x per year for CD4 testing. This is in addition to viral load and CD4 

tests that are used for disease staging and therapy initiation. Additional tests include antenatal 

screening of the 125 million births per year worldwide, testing for a panel that at least includes 

HIV and syphilis. Further, there are confirmation tests and co-infection panels that are either 

current standard of care or increasingly part of the standard in the emerging markets of the 

world. The market for this family of HIV tests is shown in the following table, indicating a nearly 

$1 billion market for POC HIV testing in the near future.

Commercialization Plan Page 52

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
US and Europe Rest of World 2017

Total POC POC Total POC POC total

Units* Share ASP Units* Share ASP Market*

Clinical HIV 

screening 60.0 20% $ 120.0 70% $ 

Antenatal screening 10.0 5% $ 120.0 70% $ $ 

Confirmation: HIV 2.0 50% $ 5.0 50% $ $ 

Co-Infection Panel 0.5 15% $ 2.0 100% $ $ 

CD4 8.0 40% $ 20.0 50% $ $ 

Viral Monitor 8.0 15% $ 60.0 50% $ $ 

$ 

*Figures in millions

In addition to infectious disease diagnostics, personalized medicine is also an attractive 

application of this technology. As pharmaceutical companies produce more specialized drugs 

that serve more narrow markets, the MBio technology will help identify potential customers at a 

reasonable cost. In 2005 the pharmaceutical industry spent over $25B in promoting new drugs. 

Prescription drug spending by consumers is projected to remain the fastest growing sector of 

health care costs. Spending on drugs is expected to account for 14.5 percent of $3.1 trillion 

health care expenditures by 2012 (for a total of $465B), compared to approximately 10 percent 

in 2001. MBio’s POCT technology could be a key tool for pharmaceutical companies to lower 

cost of patient acquisition and expand markets for their drugs. 

Other key applications for our serology and cytometry POC technologies include: 

1. Influenza Rapid Testing – The portability, low power and multi-pathogen detection 

provide an ideal tool for detection of flu strains. MBio is part of a multi-year funded 

program aimed at improved POC testing for respiratory viruses. 

2. Cardiac Trauma Testing – Cardiac panels for chest pain are an established segment in 

the POC market. Products analyze various markers in a panel, including troponin I, 

troponin T, CK-MB, BNP, and myoglobin. This market segment, for both lab and POC 

testing, has been estimated at $800 million. With the high sensitivity and multiplexing of 

the proposed technology, this is an attractive segment.

3. Systemic Inflammatory Response Syndrome (SIRS). This predecessor to sepsis kills 

over 300,000 people in the US alone. Septic shock is the most common cause of 

mortality in the intensive care unit. It is the 10th leading cause of death overall (2003) 

and is the most common cause of shock encountered by internists in the U.S. There is a 

continuum of clinical manifestations from SIRS to septic shock. Our technology can 

provide a rapid quantitative diagnostic that could diagnose the indicators of septic shock. 

4. Allergy Testing – Traditional tests are done partially at the point of care and require both 

quantitative and multi-analyte tests. It is estimated that in the allergy market alone, with 

only 1% penetration revenues could be greater than $25MM in the US and $50MM 

globally.

5. Autoimmune Disease – This very large market currently has no good diagnostic 

solutions. Evidence suggests the disease diagnosis will likely be based on simultaneous 

measurement of multiple parallel biomarkers. MBio’s multiplexed technology is uniquely 

suited for this application. 

In short, there are significant opportunities across the spectrum of medical diagnostics. 

Commercialization Plan Page 53

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Customers

We initially focus on international customers for our CD4 cytometry technology, our first product. 

Our customers are the clinics, hospitals, and healthcare systems within emerging economies; 

and international organizations and non-governmental organizations (NGO’s) that purchase and 

distribute diagnostics for the developing world. For NGO’s and system-wide purchases, the 

World Health Organization (WHO) has defined a set of criteria for the inclusion of a product into 

the bulk procurement scheme. The WHO states that for limited resource environments the 

diagnostics must operate on battery power, require no refrigeration and be simple to operate. 

Our product will meet these specifications and perform high quality tests for per test kit. 

MBio will work with its’ partners and affiliates to be included in bulk procurement schemes, both 

for specific countries as well as through umbrella organizations, such as the WHO and SCMS. 

These qualifications will add credibility and provide an objective review of our product for enduse customers. The UN procurement agencies, non-governmental organizations, the Global 

Fund to fight AIDS, TB, and Malaria, the Clinton Foundation, the PEPFAR program, and 

developing nations increasingly rely on the guidance provided by WHO on diagnostics and 

equipment that have been evaluated and intend to purchase products at reduced prices. 

The strategy also includes the global AIDS Medicines and Diagnostics Service (AMDS), which 

will ensure that poor countries have access to quality medicines and diagnostic tools at the best 

prices. The service, which will be operated by WHO, UNICEF and other partners, will help 

countries to forecast and manage supply and delivery of necessary products for the treatment 

and monitoring of AIDS. AMDS will also include a medicines and diagnostics evaluation 

component, which will ensure that manufacturers, products, procurement agencies and 

laboratories meet international quality, safety and efficacy standards. 

The customers for our CD4 product will directly overlap with customers for a 4th generation HIV 

test. Thus the channel we are currently building, through distributors and partners, will be 

ideally suited to the product envisioned in the current proposal. 

Competition

The high growth rates for POC technology have attracted a number of competitors. There are a 

number of established competitors in specific segments, such as Abbott Point of Care (glucose 

and blood chemistry), HemoCue (hematology), and Siemens (blood gas). Traditional lateral 

flow tests are marketed by Alere, Quidel, and Becton Dickinson (BD). For the segments we are 

focused on, the biggest established competitor are Alere, Trinity Biotech, and OraSure, with 

multiple brands that compete in the HIV space. The rapid tests for diagnosis of HIV marketed 

by these three competitors are all traditional lateral flow devices, and are visually read. The 

electronic readout and control features, such as procedure controls and operator lockout, 

offered by the MBio technology are valued in a professional setting, where the license of the lab 

director is on the line when point-of-care tests are not run properly. Alere is introducing a rapid 

4

th generation test that is based on lateral flow technology, but it requires a large sample (50 

microliters!) and has a complicated visual readout—virtually ensuring errors. 

In the CD4 application, Alere has introduced the PIMA CD4 counting technology for point-ofcare use. The PIMA system uses a complex pumping scheme along with immunostaining and 

fluorescence imaging to generate a CD4 cell count. The MBio system has significant 

throughput and cost advantages relative to the PIMA. We also believe that a ―fast follower‖ 

market entry is the best business model for this application. Because POC CD4 testing is a new 

concept, Alere is undertaking a number of clinical studies just to establish the operational 

characteristics of POC CD4 testing. Our market introduction will benefit from the market 

Commercialization Plan Page 54

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
development that Alere has had to do to gain acceptance for POC CD4 testing. We see an 

outstanding business opportunity with a superior new product offering that can achieve rapid 

uptake and acceptance. 

Our competitive status against select competitors is summarized below: 

Competitor Product MBio advantage

Alere Determine HIV test More sensitive and specific, lower sample 

volume, electronic readout

Trinity UniGold HIV test Higher sensitivity, electronic readout

OraSure OraQuick Higher sensitivity, electronic readout

Alere PIMA CD4 Higher throughput, simpler technology 

(more robust)

Zyomyx CD4 test Simpler procedure, electronic readout

Daktari CD4 test Lower cost, higher throughput

Competition in the point of care diagnostic market includes higher speed realization of traditional 

diagnostic approaches, ranging from rapid delivery of samples to hospital labs and third party 

diagnostic service companies such as Quest Diagnostics and LabCorp. The importance of 

POC diagnostics is pointed out by Quest Diagnostics in the 2006 annual report: ―The diagnostic 

testing industry is faced with changing technology and new product introductions. Advances in 

technology may lead to the development of more cost-effective tests that can be performed 

outside of a commercial clinical laboratory such as (1) point-of-care tests that can be performed 

by physicians in their offices and (2) home testing that can be performed by patients or by 

physicians in their offices. Development of such technology and its use by our customers would 

reduce the demand for our laboratory testing services and negatively impact our net revenues.‖ 

The clear validation of this concern is the purchase by Quest of HemoCue in 2007. 

D. Intellectual Property (IP) Protection 

There are three major pillars of our intellectual property strategy. First, we must protect our 

ideas and methods; we combine an exclusively licensed portfolio with new IP filings to provide 

protection. Second, we must establish freedom to operate for our products; we have the 

presumption of freedom to operate through our issued patents, and will supplement this with 

further analysis and licensing of any necessary IP. Finally, we will investigate the blocking

ability of our IP, whereby we could extract royalties from infringing products on the market. 

We have exclusively licensed an extensive patent portfolio from for the 

use of waveguides for making biosensors and diagnostic tests. This portfolio provides the base 

of our IP protection: (1) the portfolio provides basic coverage of the waveguide illumination 

technique, (2) the technology has been commercialized previously, showing that it can pass the 

FDA and withstand scrutiny of freedom to operate, and (3) the patents range in date from 

to the present, giving both assurance of precedence and of ongoing protection. In addition, the 

company has filed multiple applications with improvements to the waveguide technique; recently 

we have received two Notices of Allowance on filed utility patents. We will continue to 

prosecute new innovations around the platform, including a filing that covers innovation in this 

proposal. New innovation continues in sample preparation, signal enhancement, electronic 

connectivity, new assays, and new biomarkers. We have dedicated one staff member to IP 

prosecution, and have a significant budget for IP development. 

Commercialization Plan Page 55

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Protecting IP also includes managing disclosure of our ideas. We require our employees, 

consultants, outside collaborators and other advisors to execute confidentiality agreements 

upon the commencement of employment or consulting relationships with us. These agreements 

provide that all confidential information developed by or made known to the individual during the 

course of the individual’s relationship with with the company, is to be kept confidential and not 

disclosed to third parties except in specific circumstances. In the case of employees, the 

agreements provide that all inventions conceived by the individual during his or her tenure are 

the exclusive property of the company. Documents are reviewed for confidential information 

prior to public disclosure. 

Establishing absolute freedom to operate is a challenging task, as it requires complete 

knowledge of all IP in existence. In practice, the existence of patents covering one’s technology 

brings a presumption of validity of those patents, and consequently the right to manufacture and 

sell those products. This was one of the key reasons we were pleased to secure an exclusive 

license to the portfolio of patents covering the work of , 

a leader in the field of biosensors. We have the freedom to operate on these patents. 

Nonetheless, we have had discussions with potential partners about formally assessing the 

freedom to operate in the field of waveguide sensors. The relevant fields of concern are 

waveguide sensors, fluorescence immunoassays, and diagnostic microarrays. We are 

continuing to evaluate potential conflicts, and are budgeting over the 

next year for analysis and up to two freedom-to-operate opinions to clarify our position for a 

partnership. 

In addition to IP on the platform, we must secure licenses to and/or supply agreements for the 

reagents used in our tests. We anticipate being able to achieve this on commercially 

acceptable terms, and it will be a major driver for securing partnerships. 

E. Finance Plan 

MBio funding history was discussed above, and the company has already secured the funding 

for gaining initial regulatory approvals and launching first products. Moving forward the 

company will use a combination of federal funding, equity investment, and partnerships. The 

equity investment requirements are modest. While we have the cash on hand to gain approvals 

for the first products, we plan to raise an additional to complete commercialization of 

the MBio technology over the next 18 months. Funding will be used to complete the 

development of the product, secure further intellectual property protection, scale up 

manufacturing, and develop marketing relationships to promote and to distribute the product. 

The revenue generated by our technology can come from: 

1) Sales of the instrument and test kits by MBio 

2) Royalties from the sales through strategic partners 

3) Licensing or sale of the Intellectual Property to a strategic partner 

Under the first scenario MBio would retain all of the rights to the technology and develop 

manufacturing and distribution relationships to get the technology to market. This strategy is

feasible with equity fundraising, from investors and/or partners. Distribution would initially be 

through third party agents.

In the second scenario, MBio would outsource the sales and marketing to a larger distribution 

partner while managing and overseeing the design and manufacturing processes. Under this 

scenario MBio would need to increase resources in its’ manufacturing areas and engineering 

Commercialization Plan Page 56

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
capabilities and capacity. Development funding and working capital would be provided by the 

distribution partner. These resources would ensure that MBio had the ability to mass-produce 

(with contract manufacturing partners) the instrument and cartridges, with QA/QC done inhouse. 

The last scenario would be to grant an exclusive license for manufacturing and distribution of 

the products or other variation of the products. MBio has entered discussions with large 

diagnostics companies. MBio would work closely with the acquirer of the Intellectual Property to 

ensure the highest probability of commercial success. 

A combination of the above sources of revenue could develop. For example MBio could grant 

an exclusive license for the manufacturing and distribution of the product in South America while 

producing the product for distribution to the rest of the world. Further, we are considering 

licenses for certain product areas where we don’t intend to develop products.

F. Production and Marketing Plan

Operations Plan 

MBio is preparing to launch a CD4 test, and the requirements for a 4th generation HIV test would 

be similar. The company is undertaking the following significant development actions. 

 Regulatory compliance: Engineering development of a medical device must occur under an 

established quality system that meets ISO 13485 and/or FDA QSR requirements, and must 

be documented in a Design History File (DHF) that is submitted to the FDA for regulatory 

approval. A Device Master Record (DMR) is also required that fully documents the bill of 

materials, approved suppliers, all hardware and software design specifications, and all 

manufacturing procedures. Additionally, to assess and mitigate potential harm to the user 

and patient, risk management efforts incorporating human factors engineering must be fully 

documented throughout the product lifecycle. The quality system is currently being 

implemented, and the design history file for the CD4 product is actively being assembled. 

Any product resulting from the proposed development would benefit from the quality system 

currently being implemented. 

 Design for manufacture/value engineering: For viable product, material and labor costs 

must be substantially reduced from those of the prototype. Detailed engineering studies 

need to be performed to determine less expensive options for materials, components, and 

assembly methods. For the instrument, this primarily involves transitioning from expensive 

machined components to injection-molded parts, and re-designing the electronics boards 

and optical components to best balance performance with cost. For the disposable 

cartridges, there are two areas for value engineering: (1) tooling for more efficient injection 

molding, and (2) reducing labor time. The company is investing in mold tooling to lower 

cost, and investigating automation to reduce labor time and increase capacity. Validating 

laser welding of the cartridge and validating multi-cavity mold tooling are the primary 

objectives for moving the cartridge to high capacity production. 

 Supply chain management: Ramping to projected volumes will require close partnerships 

with a variety of outside contract manufacturers (CM’s) that have been approved under 

MBio’s Supplier Quality Assurance procedures. With regards to instrument production, the 

modest volumes (1000 to 10,000 per year) can easily be produced by one of many CM’s 

that have in place the required quality system and assembly infrastructure. During 

development, MBio will evaluate and qualify several CM’s, choosing one for instrument 

production. On the other hand, the much higher anticipated volumes for the CD4 

disposable cartridge, up to 20 million units per year, present significant challenges to 

production capacity. Initial production, up to 2 million units per year, is being implemented 

Commercialization Plan Page 57

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
in a new facility that MBio has leased. As the product scales up, we will investigate CM’s for 

the cartridge. MBio is currently vetting possible CM’s to do this specialized high capacity 

production, such as 

Marketing Plan 

We will roll out our products through a combination of direct sales, strategic partnerships and 

independent distributors. Our marketing strategy is to raise awareness through a full array of 

marketing activities, which include trade shows, print advertising, special programs and 

distributor promotions, to support sales in each target market. 

We will market diagnostic products directly to customers in the public health market for HIV 

testing and therapy management. This market consists of a broad range of clinics and hospitals 

and includes countries, ministries of health, states, and specific agencies such as the Centers 

for Disease Control and Prevention (―CDC‖) in the United States. We anticipate that there will be 

increasingly centralized purchasing for devices and tests for world health; as an example, the 

Global Fund is looking to centralize purchasing for the various programs it funds. There are 

also a number of organizations in the public health market, such as AIDS service organizations 

and various community-based organizations set up primarily for the purpose of encouraging and 

enabling HIV testing. We plan to register our diagnostics in a number of countries, progressing 

country by country in order of market size. We will also enlist the help of NGOs to raise 

awareness of these tests and facilitate acceptance. 

Commercialization Plan Page 58

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
G. Revenue Stream 

MBio has developed an extensive financial model for our products, both in the research use 

only markets, and the clinical diagnostics market. In addition, we have revenue from R&D 

contracts, such as the current proposal. We group the R&D contracts and our ―research use 

only‖ product revenues into a Life Sciences business line. We group our clinical diagnostics 

products, such as the CD4 product and a 4th generation HIV test, into a Clinical Diagnostics line 

of business. We anticipate introducing our CD4 product in with country roll-outs through 

 We anticipate introducing our HIV/syphilis serology product into world markets starting in 

 with country-level introductions extending over several years. The proposed HIV 4th

generation test technology would likely be introduced into world markets in and in the US 

in Additional indications, not discussed here, are included in the model. We assume 3rd

party distribution for all these products, and manufacturing at MBio’s new facility in Boulder, CO. 

Not included in this are licensing fees and royalties, which are possible (and even likely), but we 

are establishing a baseline through this model. Based on these expectations, we predict the 

following revenue for MBio, expressed in millions of $: 

 

Commercialization Plan Page 59

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
SBIR/STTR Information 

Question 8. 

MBio Diagnostics, Inc. has not received more than 15 SBIR Phase II awards from the 

Federal Government during the preceding five fiscal years. 

Prior SBIR Phase II Awards Page 60

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
PHS 398 Cover Page Supplement

1. Project Director / Principal Investigator (PD/PI)

* Title:

* Street1:

* City:

* Country:

Street2:

County/Parish:

* State:

* Zip / Postal Code:

Prefix: * First Name:

Middle Name:

* Last Name:

Suffix:

2. Human Subjects

Clinical Trial?

* Agency-Defined Phase III Clinical Trial?

* First Name:

Middle Name:

* Last Name:

Suffix:

3. Applicant Organization Contact

Person to be contacted on matters involving this application

* Phone Number: Fax Number:

Email:

OMB Number: 0925-0001 

Province:

Prefix:

Dr. Michael 

J

Lochhead

No Yes

No Yes

Cynthia

Moore

Director of Finance and Administration

5603 Arapahoe Avenue, Suite 1

Boulder

USA: UNITED STATES

CO: Colorado

80303-1377

	


 		

 !"  #

Clinical Trial & HESC Page 61

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
4. Human Embryonic Stem Cells

* Does the proposed project involve human embryonic stem cells?

If the proposed project involves human embryonic stem cells, list below the registration number of the 

specific cell line(s) from the following list: http://stemcells.nih.gov/research/registry/. Or, if a specific 

stem cell line cannot be referenced at this time, please check the box indicating that one from the 

registry will be used: 

Specific stem cell line cannot be referenced at this time. One from the registry will be used.

PHS 398 Cover Page Supplement

No Yes

Cell Line(s):

	


 		

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Clinical Trial & HESC Page 62

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
PHS 398 Research Plan

1. Application Type:

From SF 424 (R&R) Cover Page. The response provided on that page, regarding the type of application being submitted, is repeated for your 

reference, as you attach the appropriate sections of the Research Plan.

*Type of Application:

OMB Number: 0925-0001 

2. Specific Aims

3. *Research Strategy

6. Protection of Human Subjects

7. Inclusion of Women and Minorities

8. Targeted/Planned Enrollment Table

9. Inclusion of Children

10. Vertebrate Animals

13. Consortium/Contractual Arrangements

14. Letters of Support

15. Resource Sharing Plan(s)

16. Appendix

1. Introduction to Application 

(for RESUBMISSION or REVISION only)

Human Subjects Sections

2. Research Plan Attachments: 

 Please attach applicable sections of the research plan, below.

Other Research Plan Sections

5. Progress Report Publication List

11. Select Agent Research

12. Multiple PD/PI Leadership Plan

4. Inclusion Enrollment Report

New Resubmission Renewal Continuation Revision

1245-Introduction.pdf Add Attachment Delete Attachment View Attachment

1246-Specific Aims.pdf Add Attachment Delete Attachment View Attachment

1247-Research Strategy.pdf Add Attachment Delete Attachment View Attachment

Add Attachment Delete Attachment View Attachment

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1248-p24 Ag SBIR Protection Add Attachment Delete Attachment View Attachment

1249-p24 Ag SBIR Inclusion Add Attachment Delete Attachment View Attachment

1250-Targeted Enrollment.pd Add Attachment Delete Attachment View Attachment

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List of Research Plan Attachments Page 63

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
INTRODUCTION 

Here we provide an overview of revisions included in this grant resubmission. According the Summary 

Statement for the original submission, “there was significant enthusiasm for this Phase II application.” Cited 

strengths included the success of Phase I studies, promising preliminary data, and the significance of pointof-care (POC) technology. Cited weaknesses were minor. Here we attempt to address all weaknesses in 

order of appearance in the Summary Statement. Reviewer comments are in bold, followed by the MBio 

response. Italic items highlight where edits have been made to the Research Strategy. 

Not as sensitive as PCR. PCR is definitely more sensitive than antigen-based approaches, including the 

approach outlined here. We believe, however, that the cost and ease-of-use advantages of immunoassays 

will continue to result in high impact diagnostic tests, particularly at the point-of-care in limited resource 

settings. Very low cost POC PCR is still years away (in our opinion). Cost and ease-of-use are addressed 

more clearly in the Significance section. 

Uses serum or whole blood not saliva which may be more difficult. Finger stick whole blood is widely 

accepted worldwide and will be necessary for the antigen detection part of the proposed assay. Oral fluid is 

very convenient, but antigen detection in saliva is not likely to meet analytical sensitivity requirements. 

Clinical assays are available to detect HIV-1 antibody/antigens. Clinical lab assays indeed exist (e.g., 

Abbott and Bio-Rad). But high quality POC antigen/antibody combination assays have still not been well 

established. Published data for the antigen detection component of the Alere Determine® HIV-1 Ag/Ab 

Combo assay (e.g., recent posters from CROI 2013, Atlanta) show significant opportunity for improvement, 

particularly in the context of acute infection detection. 

POC kits based on nucleic acid based testing for early HIV infection may be a more appropriate 

technology. Nucleic acid tests perhaps pose the greatest commercial threat to the proposed antigenbased system. As discussed above however, we believe the cost advantages and ease-of-use of 

immunoassay approaches will keep antigen detection very competitive in cost-driven applications such HIV 

screening in global markets. Cost and ease-of-use are addressed more clearly in the Significance section.

No details are given for VDSA (trade secret). A detailed description of the viral disruption formulation is 

not critical for the scientific review of this proposal. As the reviewer correctly notes, viral disruption and 

immune complex disruption have “standard” elements, and the details of the formulation are not at the core 

of our innovation claim. Instead, our innovation is around integration of the disruption approaches into the 

POC system. 

It is not clear what was innovative in these efforts. We believe the combination of low cost, quantitation, 

and sensitivity is unique in this proposal. We try to more clearly articulate this in the Innovation section.

Of few quantitative results presented, judging from figure 3 it is not convincing whether the results 

support the claimed analytic sensitivity and detection limit. We articulate our method of defining 

reactivity in Fig. 3, where we run the no target control samples in triplicate and define cutoff as mean plus 5 

standard deviations. We stand by this definition. Regardless, the performance of the particle assay is not 

where we wanted it to be for making the transition to product, and we clearly stated that the “One-Step 

protocol does not meet the target analytical sensitivity milestone.” It was this assessment that led us to 

develop the direct waveguide sandwich assay, which yielded the Table 1 performance. The direct 

waveguide assay is at the center of the proposed Phase II research. 

Why are there huge reading variations or errors at higher concentrations and almost no variation or 

error at lower concentration? Is it because the signal is too low to produce any reading variation? 

Data are shown on a linear plot which compresses the appearance of relative variation. The variation, 

however, is high across the entire dynamic range and is concentration independent. The high variability 

makes the particle approach less attractive in general. This was part of the justification for our proposed 

transition to the direct waveguide sandwich immunoassay approach. This point is augmented in the 

resubmission. 

Introduction Page 64

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
SPECIFIC AIMS 

HIV infection remains a major public health crisis both in the United States and worldwide. There is 

increasing awareness that acutely infected individuals disproportionately contribute to disease spread (1). 

Yet these individuals remain the most difficult to identify, as infectivity is highest prior to the appearance of 

the HIV antibodies that serve as the basis for serological diagnostics (2). There are currently no FDAapproved point-of-care (POC) tests that are sensitive to acutely infected individuals. An HIV-1/2 

antigen/antibody (Ag/Ab) combination assay – the so-called “4th generation” immunoassay – in an 

inexpensive, simple to use, POC format would fundamentally improve HIV-1/2 screening efforts in the 

United States and worldwide (3, 4). 

MBio Diagnostics, Inc. is developing a point-of-care infectious disease testing platform for multiplexed HIV 

and coinfection serodiagnostic screening. Prototype devices have been placed in field sites in San Diego, 

Mozambique, Kenya, and Brazil. Due to cost and labor constraints, current acute infection diagnosis is 

typically based on pooled sample nucleic acid amplification testing algorithms, with 7 to 14 day turnaround 

times. 4th gen Ag/Ab assays in the clinical laboratory have been approved recently (Abbott ARCHITECT 

HIV Ag/Ab combo, Bio-Rad GS HIV 1/2 Ag/Ab Combo), but the 4th gen clinical analyzers do not offer the 

improved linkage to care associated with rapid, POC HIV testing. Here we propose an inexpensive device 

that delivers the performance of lab-based 4th gen Ag/Ab combo assays in a simple, POC package. We 

build on successes of our Phase I SBIR program and propose continuation of our translational research on 

a novel system with high commercial potential, offering:

 Parallel HIV-1/2 antibody and p24 antigen detection on a single point-of-care platform. 

 Workflow and ease-of-use comparable to conventional HIV rapid tests. 

 Robust, low cost, minimally instrumented system for use in emergency departments, public health 

labs, STD clinics, and targeted outreach programs. 

Aim 1: Assay Development. Combine the Phase I p24 antigen detection assay with the MBio multiplexed 

serology assay cartridge, addressing issues of final monoclonal antibody (mAb) pair selection, HIV-1 Ag 

and HIV-2 Ag selection, reagent conjugations, cross-reactivity, and minimization of assay steps and 

complexity. The Aim 1 milestone is an HIV-1/2 antigen/antibody detection assay with performance 

equivalent to FDA-approved laboratory 4th gen Ag/Ab combo assays for the MBio early/acute sample 

collection [a set of 5 commercially available HIV-1 seroconversion/performance panels, two anti-HIV-1/2 

combo performance panels, an anti-HIV-2 performance panel, and a unique collection of acute samples 

from San Diego.]

Aim 2: Cartridge Integration. Modify the MBio Cartridge, Rack, Reader, and Software to deliver an 

automated HIV-1/2 Ag/Ab combo result, and incorporate heat stable assay reagents into the MQ 

cartridge. The Aim 2 milestone is a portable, integrated system delivered to clinical collaborators that 

meets FDA CLIA waiver guidance requirements. 

Aim 3: Assay Validation. Validate system using well characterized early HIV infection specimens 

including a panel of 200 HIV positive specimens comprised of 20 acute infection samples (RNA+ / Ab -), 

early seroconversion (Western Blot indeterminate) and seropositive (HIV-1 and HIV-2) samples. 200

HIV-negative samples will be used for specificity testing. The Aim 3 milestone is a dataset 

demonstrating performance equivalent to FDA-approved laboratory 4th gen HIV-1/2 Ag/Ab assays. 

Aim 4: Pre-Market Field Evaluation. Place systems in intended use setting and capture operational and 

usability feedback in advance of design lock; and generate a preliminary dataset on capillary whole 

blood samples from 100 study participants in San Diego. The Aim 3 and 4 milestone is a system design 

and dataset for an FDA investigational device exemption (IDE) meeting in advance of clinical trials.

The assembled group of investigators is uniquely capable of executing this project in a timely and cost 

efficient manner. The PI, Michael Lochhead, Ph.D. has led successful life science product 

commercialization efforts and manages MBio R&D programs, including several NIH grants projects. The 

MBio team includes established diagnostics industry veterans, development engineers, and bioassay 

scientists. Complementing the MBio group is a world class clinical team at the University of California, San 

Diego, with a well-established early HIV infection research program. 

Specific Aims Page 65

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
[Edits to the original submission are included in bold brackets in this resubmission] 

SIGNIFICANCE 

HIV/AIDS remains a critical public health crisis in the United States and worldwide: 

 CDC estimates there are 1.2 million Americans with HIV, and 1 in 5 do not know their disease status (1). 

 Because the early/acute phase of infection is marked by very high titers of active viral particles (>3E05 

particles/mL), infectivity is significantly higher for individuals during the early/acute phase of infection 

compared to those with chronic HIV infection and a mature antibody response. 

 New guidelines from CDC for laboratory testing of HIV infection recommend initial screening with a 

sensitive, ―4th generation‖ antigen/antibody combination assay (2). 

 The global impact of HIV/AIDS remains enormous, with approximately 2.7 million new infections per year, 

2 million AIDS-related deaths, and ~33 million people living with HIV (3). 

Current point-of-care HIV testing in the United States is based on FDAapproved rapid tests. Although these antibody-based tests have 

excellent clinical sensitivity and specificity for seroconverted individuals, 

they do not identify pre-seroconversion, acutely infected individuals (4, 

5). Recent studies have demonstrated that measurement of HIV p24 

antigen in combination with antibody measurements can significantly 

improve identification of acute infection cases (5). 4

th generation HIV-1/2 

Ag/Ab combo assays on clinical analyzer instruments have been 

commercially available outside of the United States for several years. 

But these 4th gen systems, such as the Abbott ARCHITECT® (FDA 

approval issued June 2010), are large clinical instruments not designed 

for point-of-care use. Inverness Medical recently launched a point-ofcare, lateral flow based p24/antibody combo assay (Determine®

 HIV-1 

Ag/Ab Combo, not currently approved in the US). Recent results 

suggest that the antigen feature of the Determine® test provides an 

advantage over antibody-only HIV-1/2 rapid tests, but that the antigen 

sensitivity is inferior to the clinical analyzers (6-9), and the specificity of the antigen line is a concern (10). 

PCR-based methods deliver the sensitivity required for acute infection diagnosis, but current PCR-based 

molecular tests do not meet the cost, turnaround time, or ease-of-use requirements needed for the largescale public health screening. 

Effective ELISA tests for the HIV-1 p24 antigen have been available for some time, but are not approved for 

human diagnostics. Importantly, the ELISA protocol includes pre-treatment to break antigen-antibody (AgAb) complex, long incubations, various wash protocols, and added cost that make this test format 

incompatible with sensitive POC testing. The system proposed here avoids the Ag-Ab complex issue by 

reporting parallel Ag and Ab results. Other sensitive p24 assays have appeared in the literature recently 

(11), including POC devices with clinically relevant sensitivities and workflow (12, 13). The POC systems 

are promising, but they require multiple steps and, because they are built on lateral flow technology, those 

systems will not have the multiplexing capabilities of the system described in this proposal. 

[While PCR and lab-based methods provide outstanding sensitivity, they will be limited in impact in 

high disease burden, resource-limited settings where cost and ease-of-use are major drivers. The 

system proposed here addresses a major unmet public health screening need.]

INNOVATION 

A robust, POC device with HIV Ag/Ab with performance equivalent to laboratory testing would significantly 

advance HIV screening efforts in the United States and globally by identifying the most infectious individuals 

quickly and cost effectively. The research proposed here has the ultimate goal of delivering direct detection 

of HIV-1/2 viral antigen at picogram/ml sensitivity, and HIV-1/2 antibody reactivity, from fingerstick whole 

blood in a simple point-of-care device. In order to do this, a highly innovative cartridge-based 

immunofluorescence assay approach is proposed. The major accomplishment of Phase I is a substantial 

Figure 1. MBio multiplexed 

immunoassay system, including a 

reader and disposable cartridges.

Research Strategy Page 66

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
dataset on the MBio system demonstrating equivalent performance to significantly more complex laboratory 

tests – feasibility is clearly established. The major innovation of Phase II will be integration of the simple, 

finger stick whole-blood assay into the cartridge and reader system capable of operation in non-CLIA 

laboratories. Performance is achieved through a highly innovative combination of fluidic cartridge design, 

optical instrumentation, viral lysis techniques, and (if needed) an integrated wash mechanism. The 

disposable cartridge is assembled from injection-molded plastics with feature sizes within well-established 

manufacturing tolerances. The cartridge incorporates a molded waveguide that enables sensitive 

fluorescence immunoassays without the fluid handling requirements of ELISAs or the alignment, pumping, 

or interface issues of microfluidic devices—and the innovation of the key features of this waveguide 

technology were recently recognized by the patent office, with a Notice of Allowance for a patent on the 

technology. MBio’s technical innovations are directed at simplicity of the user experience (minimal 

steps, small sample, error tolerance, heat-stable on-board reagents, etc) while not compromising on 

performance. Cartridge assembly costs will be comparable to those of existing lateral flow rapid tests, 

which have well established < $1 per piece manufactured cost. The novel, waveguide-based CartridgeReader System solves the light coupling and reproducibility issues that have plagued waveguide based 

fluorescence sensors for decades. 

To our knowledge, there is no POC assay system on the horizon in the United States that can deliver the 

performance and ease-of-use expected from the MBio system. NIAID funding will provide the critical 

resources required to complete development, positioning the company to justify investment in clinical trials 

and manufacturing scale-up. Phase II will position the company for FDA pre-IDE meetings, and will provide 

the groundwork for pre-market approval (PMA) clinical trials and subsequent CLIA waiver evaluations. 

APPROACH 

Preliminary Studies and Phase I Final Report 

 The overall goal of Phase I was the development of a highly 

sensitive HIV p24 antigen assay, with post-Phase I efforts 

leading to an HIV-1/2 Ag/Ab combination assay in an 

inexpensive, simple to use, point-of-care (POC) format. To 

achieve high p24 Ag detection sensitivity, a multi-particle 

immunoassay sandwich approach was proposed (Fig. 2). With 

capture antibody conjugated to a paramagnetic particle, and 

detect antibody conjugated to an intensely fluorescent particle, 

the feasibility of combining target antigen concentration and 

labeling was demonstrated in the preliminary studies. HIV-1/2 

Ag/Ab combination assay development, via integration of the 

developed multi-particle p24 Ag assay with the MBio HIV-1 

antibody assay plus incorporation of HIV-2 antibody detection, 

was slated for Phase II. 

Phase I, Specific Aim 1. Establish a core collection of well-characterized clinical serum/plasma samples 

focused on acute and recent HIV infection. The Phase I collection will be based on samples from existing 

archives and sources, with additional testing under this award. The Phase I milestone will be a collection of 

60 HIV-1 positive and 40 negative sera. 

The Aim 1 milestone was achieved. A collection of 40 HIV-negative serum samples was obtained from 

Valley Biomedical (Winchester, VA). The following HIV-1 seroconversion and performance panels, each 

including challenging acute/early HIV-1 infection samples and/or low anti-HIV-1 titer samples, were 

acquired: seroconversion panels HIV 9075, HIV 9079, and HIV 12008 from ZeptoMetrix (Buffalo, NY), and 

seroconversion panel PRB965 and anti-HIV-1 low titer performance panel PRB109 from SeraCare Life 

Sciences (Milford, MA). Added to this set were two HIV-1 positive control serum samples (SeraCare).

Figure 2. Particle complex approach to 

sandwich immunoassays. The magnetic 

particle provides concentration and specific 

signal enhancement. The fluorescent 

particle provides signal amplification.

Research Strategy Page 67

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
In addition to these commercially available samples, we assembled a panel of early/acute samples from our 

clinical collaborators in San Diego. For Phase I we obtained five acute (HIV RNA+/EIA-) plasma samples 

and 10 HIV RNA-positive plasma samples that yield indeterminate Western blot band patterns (Dr. Susan 

Little, UCSD Antiviral Research Center). 

Phase I, Specific Aim 2. Demonstrate MBio’s magnetic multi-particle approach for rapid, ultra-sensitive, noenzyme detection of HIV p24 antigen. The milestone for this aim is a model assay standard curve for p24 

antigen detection with a lower limit of detection of 20 pg/ml. 

The Aim 2 multi-particle assay concept was demonstrated using the following key reagents: capture antip24 mAb conjugated to paramagnetic beads, biotinylated detect anti-p24 mAb, and NeutrAvidin (NA)-

coated fluorescent beads. The start of Phase I used the following preliminary assay protocol: the antibodyconjugated magnetic beads were combined with the sample and biotinylated mAb then incubated for 25 min 

at ambient temperature. The magnetic beads where ―pelleted‖ with a magnetic separator, the supernatant 

was discarded. The magnetic beads were then suspended in buffer containing the NA-coated fluorescent 

beads and incubated for 10 min. Following a wash step, this reaction mixture was transferred to a customdesigned disposable cartridge and magnetic particles were translocated with a magnet to separate 

magnetic bead:fluorescent bead complexes from free fluorescent beads. The cartridge was imaged on a 

fluorescence microscope and image analysis software was used to generate fluorescent bead counts. 

Phase I goals were to optimize the assay reagents and to simplify this preliminary protocol. A range of 

magnetic particle types, differing in size and/or surface functionality (carboxylic acid, amine, N,N‘-

carbonyldiimidazole) where evaluated. Coupling to carboxylic acid MyOneTM particles via sulfo-NHS/EDC 

chemistry provided the highest yields of conjugated active mAb. Dark red fluorescent particles (Thermo 

Scientific) of varying sizes and surface functionalities were tested. The best assay performance achieved 

to-date has been with NeutrAvidin adhered to 0.39 m sulfate-functionalized fluorescent particles. 

A set of candidate anti-p24 antibodies, consisting of three polyclonal antibodies and 13 monoclonal 

antibodies, were screened for multi-particle assay performance (protocol as outlined above). Although the 

assay protocol as outlined above performed well with p24 Ag spiked into buffer, serum samples (50% v/v in 

assembled bead binding reactions) initially posed problems: significant aggregation of the magnetic beads 

and adherence of magnetic beads to the cartridge surface. Additionally, certain HIV-negative serum 

samples yielded false-positive results. These were subsequently identified as samples containing 

heterophilic antibodies (HAs). Both problems were alleviated through assay buffer optimization and the 

introduction of an HA competitor. 

“Two-Step” Multi-Particle Assay. The following optimized ―Two-Step‖ protocol was utilized for p24 Ag low 

limit of detection (LLOD) determination, for calculating the %CV at the limits of detection, and for evaluating 

assay performance with the Aim 1 sample set. Complete assay protocol details are not included here due to 

space constraints. Briefly, 25 μl of serum or plasma was added to reagent buffer containing both the the 

mAb-magnetic particles and biotinylated detection mAb. Following a 30 min room temp incubation, a 

magnetic separator was used to collect the particles, which were then resuspended in NeutrAvidin-coated 

fluorescent particles. Following a brief incubation, a magnetic wash-resuspension was used to remove free 

fluorescent particles. Particles were then magnetically focused and imaged on a fluorescence microscope 

and custom image analysis software provided the number of magnetic bead-fluorescent bead complexes. 

We note here that we use the World Health Organization International Standard HIV-1 p24 reagent (NIBSC 

code: 90/636), expressed in international units (IU/mL). The NIBSC reference reagent is commonly used 

for cross-platform analytical comparisons (14), and it serves as the basis for analytical sensitivity 

performance standards from the European Commission; HIV-1 antigen assays used for diagnostics 

purposes must have an analytical sensitivity of 2 IU/mL or better using the NIBSC standard. 

The lower limit of p24 Ag detection with the Two-Step multi-particle protocol was determined via serial 

dilutions. As shown in Fig. 3(a), the limit of detection with this protocol is approximately 1.2 IU/mL p24 Ag. 

This compares well with the limits of detection of this same p24 reference standard measured with four 

Research Strategy Page 68

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
commercial HIV Ag/Ab combination assays and one HIV p24 Ag only assay: ARCHITECT HIV Ag/Ab 

Combo (1.24 IU/ml), AxSYM HIV Ag/Ab Combo (1.94-2.25 IU/ml), VIDAS HIV DUO Quick (0.43 IU/ml), 

VIDAS HIV DUO Ultra (0.66 IU/ml), and VIDAS HIV p24 II (0.73-1.15 IU/ml) (14). 

Having delivered a multi-particle assay with analytical sensitivity approaching the Abbott ARCHITECT (2 

IU/mL versus 1.24 IU/mL) is a major accomplishment of Phase I Aim 2. As the Fig. 3 standard deviation 

bars suggest, however, assay variability is higher than desired. An additional Aim 2 goal was to simplify the 

protocol, reducing the assay time to ≤ 30 min and the number of steps to no more than two requiring user 

interactions. Efforts shifted to protocol minimization and improved reproducibility. 

“One-Step” Multi-Particle Assay. The Two-Step protocol has the kinetic advantage of an extremely high 

affinity biotin-NA interaction bringing the magnetic beads, with bound p24 Ag immunocomplexes, and 

fluorescent beads together. The tradeoff, however, is a requirement for the separate addition of NA-coated 

fluorescent beads and a second binding reaction incubation. To achieve the minimal protocol as outlined in 

Fig. 2, the preparation of fluorescent beads pre-bound/conjugated to detect antibody was necessary. 

Extensive optimization of chemistry, bead ratios, incubation times, etc. resulted in the following simplified 

protocol: A 25-l sample is added to a reagent buffer containing mAb-magnetic particles and mAbfluorescent particles (see Fig. 2). Following a 30-min incubation at room temperature with rotational mixing, 

a magnetic wash is performed, particles are magnetically focused, and then analyzed as described above. 

The lower limit of p24 Ag detection with the ―One-Step‖ multi-particle protocol was determined by assaying 

12.5, 25, 50, 100, and 200 IU/ml WHO Standard p24 Ag spiked into normal serum. Each concentration was 

assayed in triplicate. An intra-experiment cutoff value [mean + (5 X SD)] was calculated from results 

obtained with five replicates of a no-spike normal serum control. As shown in Fig. 3(b), the LLOD is 

approximately 30 to 40 IU/mL, a 20-fold higher concentration than achieved with the Two-Step protocol. 

Unlike with the Two-Step protocol, the number of detected p24-dependent bead pairs increased almost 

linearly with incubation periods beyond 30 min (1 hr and 2 hr tested; data not shown), demonstrating the 

slow binding kinetics between these two relatively large particles. The One-Step protocol does not meet the 

target analytical sensitivity milestone. 

Direct Waveguide Sandwich Immunoassay. Concurrent with multi-particle assay development, MBio also 

developed a direct p24 Ag sandwich assay on MBio‘s SnapEsiTM planar waveguide platform under Aim 2. 

The sandwich assay was initially a reference method for particle assay development, but improvements to 

the direct sandwich assay approach progressed to the point where it was out-performing the multi-particle 

assays both in terms of sensitivity and simplicity. Fig. 4 provides a schematic cross-section of the MBio 

planar waveguide cartridge (15). The sandwich assay protocol is discussed in more detail below. 

Figure 3. (a) Two-Step Multi-Particle Assay analytical sensitivity. (b) One-Step Multi-Particle Assay analytical 

sensitivity. NIBSC p24 antigen reference standard spiked into normal serum. Dashed line is cutoff, defined as mean 

signal + 5 standard deviations of 8 replicate measurements of the normal serum zero. Error bars are ± 1 stdev. (c)

Schematic of MBio multiplexed immunoassay cartridge (image from reference (15)). For the direct p24 sandwich 

immunoassay, capture antibody is printed to the waveguide substrate, and detect antibody carries a label. See text.

0

500

1000

1500

2000

0 5 10 15 20

Particle Counts

p24 Conc. (IU/mL)

0

500

1000

1500

2000

2500

0 50 100 150 200 250

Particle Counts

p24 Conc. (IU/mL)

Research Strategy Page 69

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
The main conclusion of the Aim 2 effort is that although the multi-particle assay approach came 

close to achieving the stated sensitivity milestone, an alternative approach that is both simpler and 

more sensitive was discovered during assay development. The direct sandwich immunoassay using 

the MBio planar waveguide has become the focus of Phase II. 

Phase 1, Specific Aim 3. Quantitatively compare performance of the MBio antigen/antibody assay on the 

Aim 1 recent infection panel with gold-standard methods. The milestone for this aim is detection of 80% of 

HIV-1 positive samples not detected with the current market-leading FDA-approved HIV-1/2 rapid test. 

The performance of the Two-Step multi-particle p24 Ag assay, the SnapEsiTM planar waveguide p24 Ag 

sandwich immunoassay, and the SnapEsiTM planar waveguide 3rd Generation HIV-1 serology assay were 

compared with the ―recent infection‖ panel (developed under Aim 1). With each of these assays, cutoff 

values were determined by assaying 40 HIV-negative serum samples; positive assay cutoffs were 

calculated as follows: cutoff = mean + (5 X Standard Deviation). For comparative purposes, the same 

recent infection panel was assayed at MBio with the following two rapid tests and two EIA‘s: OraQuick 

Advance® Rapid HIV-1/2 Antibody Test (OraSure Technologies, Inc., Bethlehem, PA), Uni-GoldTM 

Recombigen® HIV (Trinity Biotech, PLC, Wicklow, Ireland), Alliance HIV-1 P24 Antigen ELISA (PerkinElmer, 

Waltham, MA), and GS HIV-1/2 Combo Ag/Ab EIA (Bio-Rad, Hercules, CA). The product insert protocols 

were strictly followed for each of these commercial tests. The results are provided in Table 1. 

Table 1 Notes: 

All tests were run at MBio Diagnostics, with the exception of the Abbott ARCHITECT (package insert data provided) 

(A) High Ab titer HIV-1 positive control serum samples (SeraCare) 

(B) Anti-HIV-1 low titer performance panel PRB109 (SeraCare). 

(C) Seroconversion panel PRB965 (SeraCare). 

(D) Seroconversion panel HIV 9075 (ZeptoMetrix). 

(E) Seroconversion panel HIV 9079 (ZeptoMetrix). 

(F) Seroconversion panel HIV 12008 (Zeptometrix). 

(G) Acute infection plasma samples (HIV RNA+/EIA-, Dr. Susan Little, AVRC, UCSD). 

(H) HIV RNA+ samples that yielded indeterminate Western blot band patterns (Dr. Susan Little). 

† PRB109-04 is a bleed from the same donor as PRB109-03, collected 2 days later. 

‡ PRB109-11 is a bleed from the same donor as PRB109-10, collected 2 days later. †† PRB109-13,14, and 15 are 

same donor bleeds collected 5 days and 2 days apart, respectively. 

‡‡ PRB109-18 is a bleed from the same donor as PRB109-17, collected 2 days later. 

The anti-p24 antibody screen and heterophilic antibody (HA) background reduction investigations 

completed during multi-particle assay development aided development of the waveguide direct sandwich 

assay. The same detection mAb is used for both assays and a similar HA blocking strategy is employed. 

The following protocol was used to generate the ―MBio Rapid HIV p24 Ag‖ assay results shown in Table 1: 

21 l of sample was mixed with 9 l of detect reagent (including biotin-mAb) and added to the cartridge 

entrance port. Following a 20-min room temp incubation, SA-SureLight (SLP3) was added to the cartridge 

inlet port, displacing the sample volume. After a 15-min incubation at room temperature, wash buffer was 

added to the inlet port and the cartridge was analyzed on the MBio Reader (read time ~ 30 seconds). 

MBio HIV-1/2 Antibody Assay. Although Phase I development focused primarily on p24 antigen detection, a 

useful POC 4th gen assay must combine HIV-1/2 antibody detection as well. The MBio rapid HIV-1 Ab 

assay currently under development is a 3rd generation assay that employs immobilized HIV-1 antigen gp41 

and fluorescently-labeled detect gp41; sample anti-gp41 antibodies bridge the capture and detect antigens. 

In addition to being simpler than 2nd generation assays, the 3rd generation test does not rely on the 

specificity of an anti-human IgG detect Ab and therefore has the advantage of additionally detecting antigp41 IgM antibodies in early infection samples. The protocol used to generate the ―MBio Rapid HIV-1 Ab 

(gp41)‖ assay data in Table 1 is as follows: 15 l of sample was added to a detect reagent containing 

A647-labeled gp41 and then immediately transferred to the cartridge inlet port. Following a 20-min room 

temp incubation, wash buffer was added and the cartridge was analyzed on the MBio reader. 

Several important conclusions can be drawn from the Table 1 reference method comparison 

 The MBio HIV-1 Ab assay significantly outperforms the FDA-approved rapid tests 

Research Strategy Page 70

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
 When MBio HIV-1 Ag and Ab data are considered together, the combination compares very 

well to FDA-approved laboratory 4th generation Ag/Ab Combo assays (Bio-Rad and Abbott) 

 Assay performance feasibility is established; Phase II will focus on system integration

Table 1. Summary of Results – Platform Comparison (see text for description)

 Panel Member 

I.D.

OraQuick 

Advance

Trinity

Uni-Gold

PerkinElmer 

HIV p24 Ag 

(S/CO)

Bio-Rad GS 

HIV Combo 

Ag/Ab (S/CO)

Abbott 

ARCHITECT 

Ag/Ab Combo* 

(S/CO)

MBio HIV-1 

Ab (gp41)

(S/CO)

MBio Direct 

p24 Ag

(S/CO)

MBio

Multi-Particle 

HIV p24 Ag 

(S/CO)

SC-9182257 POS POS 0.4 > 16.4 36.8 0.2 0.5

SC-9148134 POS POS 0.5 16.3 35.9 0.1 0.4

PRB109-01 NEG NEG 22.9 14.8 40.9 2.7 14.3 3.2

PRB109-02 NEG NEG 31.8 > 16.4 281.3 2.1 115.3 8.1

PRB109-03† NEG NEG 0.9 5.8 3.5 1.0 0.5 0.4

PRB109-04† NEG NEG 2.0 13.0 10.7 2.7 0.4 0.2

PRB109-05 NEG POS 0.4 14.6 10.2 14.3 0.4 0.2

PRB109-06 NEG POS 3.6 7.3 4.1 2.2 5.1 1.3

PRB109-07 NEG NEG 0.6 1.7 3.9 0.8 1.1 0.4

PRB109-08 NEG NEG 0.3 0.2 0.3 0.5 0.2 0.3

PRB109-09 NEG NEG 2.4 13.3 10.6 3.1 3.2 0.5

PRB109-10‡ NEG POS 0.6 14.6 20.5 5.7 0.2 0.2

PRB109-11‡ POS POS 1.1 15.0 36.5 5.0 1.0 0.4

PRB109-12 NEG POS 1.8 15.1 34.1 14.4 2.0 0.3

PRB109-13†† NEG NEG 2.7 10.6 23.6 0.3 0.4 0.3

PRB109-14†† NEG POS 9.6 > 16.4 127.5 5.4 0.7 0.6

PRB109-15†† POS POS 7.6 16.2 127.9 11.8 0.6 0.1

PRB109-16 NEG POS 21.2 15.7 116.7 2.5 40.8 6.8

PRB109-17‡‡ NEG NEG 4.2 15.3 41.6 1.8 11.2 0.6

PRB109-18‡‡ POS POS 3.8 14.9 58.6 7.2 1.9 0.3

PRB109-19 POS POS 0.5 15.0 23.5 9.0 0.5 0.2

PRB109-20 NEG NEG 0.8 10.7 13.7 3.4 0.1 0.2

PRB965-01 NEG NEG 0.2 0.3 0.2 0.1 0.3 0.2

PRB965-02 NEG NEG 0.5 3.2 2.8 0.2 0.6 0.5

PRB965-03 NEG NEG 0.6 3.4 3.5 0.2 1.1 0.5

PRB965-04 NEG POS 0.7 13.3 15.6 4.8 1.2 0.6

PRB965-05 POS POS 0.5 15.8 22.4 6.1 0.5 0.6

PRB965-06 POS POS 0.3 15.2 18.8 9.9 0.3 0.4

ZM 9075-02 NEG NEG 0.2 0.2 0.1 0.4 0.3 0.1

ZM 9075-03 NEG NEG 1.6 5.8 7.3 0.7 3.9 0.1

ZM 9075-04 POS POS 2.0 14.7 31.9 11.5 4.0 0.2

ZM 9075-05 POS POS 0.6 15.0 51.0 13.1 0.7 0.1

ZM 9079-08 NEG NEG 0.2 0.2 0.1 0.5 0.1 0.4

ZM 9079-09 NEG NEG 0.9 5.0 3.6 0.5 1.7 0.8

ZM 9079-10 NEG NEG 3.4 14.0 23.9 0.5 17.9 0.6

ZM 9079-11 NEG NEG 6.4 15.5 43.1 1.1 23.4 2.6

ZM 9079-12 NEG NEG 11.3 16.2 68.7 2.8 53.0 4.7

ZM 9079-13 NEG IND** 0.8 12.3 34.8 2.1 0.7 0.2

ZM 9079-14 NEG NEG 0.6 11.1 3.8 1.9 0.6 0.3

ZM 9079-15 NEG NEG 0.3 9.9 4.1 3.6 0.2 0.3

ZM 9079-16 NEG IND** 0.3 9.8 8.4 3.2 0.2 0.8

ZM 12008-06 NEG NEG 0.2 0.2 0.1 0.2 ND 0.1

ZM 12008-07 NEG NEG 0.2 0.2 0.1 0.3 0.6 0.1

ZM 12008-08 NEG NEG 0.3 0.6 0.5 0.2 0.5 0.1

ZM 12008-09 NEG NEG 7.8 15.0 50.3 0.2 13.6 3.2

ZM 12008-10 NEG NEG 28.4 16.3 167.7 0.8 57.6 8.1

ZM 12008-11 NEG NEG 11.8 14.1 53.7 6.8 2.0 0.4

ZM 12008-12 POS NEG 0.6 15.4 28.4 8.9 0.3 0.6

ZM 12008-13 POS NEG 0.3 15.7 30.2 8.9 0.4 0.1

0.0

177-SL (Acute) NEG NEG 21.0 16.1 0.3 0.1 0.4

189-SL (Acute) NEG NEG 0.8 2.6 0.6 0.2 0.4

190-SL (Acute) NEG NEG 6.5 14.2 0.4 2.8 0.6

191-SL (Acute) NEG NEG 10.4 13.7 0.4 6.7 1.0

198-SL (Acute) NEG NEG 0.3 0.3 0.9 0.0 0.2

178-SL (IND WB) NEG POS 1.2 13.8 5.2 0.3 0.2

187-SL (IND WB) POS NEG 0.7 14.4 2.1 0.1 0.2

192-SL (IND WB) NEG POS 1.0 14.8 2.1 0.3 1.0

193-SL (IND WB) NEG POS 4.5 14.2 2.4 0.7 0.5

194-SL (IND WB) POS NEG 1.0 15.4 1.8 0.9 0.4

195-SL (IND WB) NEG POS 0.5 6.4 5.5 0.0 0.3

196-SL (IND WB) POS POS 0.9 15.7 8.6 0.5 0.1

197-SL (IND WB) POS NEG 6.4 14.7 2.7 1.1 0.6

199-SL (IND WB) POS POS 0.8 16.0 7.9 0.1 0.3

200-SL (IND WB) POS POS 7.0 15.9 2.8 0.8 0.3

 * Data provided in product inserts

** Indeterminant (one observer called as negative; two observesr called as very weak positive.

F

G

H

HIV Rapid Tests Enzyme Immunoassays (EIA)

A

B

C

D

E

Research Strategy Page 71

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Of the 64 samples in Table 1, the Bio-Rad GS HIV Combo Ag/Ab EIA and Abbott ARCHITECT Ag/Ab 

Combo results indicate that 56 are HIV-positive (sample 198-SL, with an HIV RNA concentration of 

approximately 2,000 copies/ml, was missed by both assays). The MBio p24 Ag and HIV Ab serology 

combination identified 52 of these 56 as HIV-positive, a 92.9% sensitivity. The OraQuick Advance HIV 1/2 

Ab and Trinity Uni-Gold HIV-1 rapid tests identified 18 and 24 of the 56 as HIV-positive, respectively, for 

assay sensitivities of 31.1% and 42.9%. Of the HIV-positive samples not detected by the OraQuick 

Advance and Trinity Uni-Gold rapid tests, 84.2% and 83.1%, respectively, were identified as HIV-positive by 

a combination of the MBio Rapid p24 Ag and HIV-1 Ab Rapid Ab planar waveguide assays. Thus, a 

combination of the Ag/Ab planar waveguide formats fully meets the goals of the Phase I program.

Although the MBio p24 Ag assay performed very well, it is sensitive only to extraviral p24 antigen. In 

contrast, the PerkinElmer ELISA assay includes an immune complex dissociation procedure that both 

denatures sample antibodies, releasing bound p24, and disrupts virus particles, releasing additional 

detectable p24. An advantage of 4th generation Ag/Ab assays is that immune complex dissociation is not 

required—antigen and antibody detection overlap within the seroconversion window. However, we

recognized that increasing the concentration of detectable p24 via virus disruption without significantly 

compromising the serology component of a 4th generation assay would improve assay sensitivity. 

To this end, non-ionic detergents alone and in combination with ionic detergents were evaluated for 

effectiveness at improving p24 Ag detection without compromising the ability to detect sample antibodies. 

Although further optimization is pending (Phase II), a preliminary virus disruption sample additive (VDSA) 

has been developed. Details of the VDSA formulation are being held as trade secret by MBio. For the 

purposes of this grant review, we can say we are using a combination of ionic detergents, salts, and 

chelating agents. [We are not claiming details of the viral disruption formulation as a core innovation 

of this grant proposal. Instead, it is incorporation of disruption into the simple worklflow that will be 

unique.] The assay protocol was as described above for the MBio Rapid HIV p24 Ag assay with following 

exception; with the  ̳VDSA‖ protocol, assays were initiated by combining 

sample with the VDSA reagent, with mixing via aspiration. Detect 

reagent (includes biotin-mab and blockers) was then added to the 

sample-VDSA mixture and the entire mix was then transferred to the 

cartridge inlet port. A comparison of this ―virus disruption‖ protocol to the 

MBio Rapid HIV p24 Ag assay protocol described above was performed 

with the five acute HIV infection samples listed in Table 1 (G). Results 

from this comparison are shown in Table 2. By combining the MBio 

planar waveguide Ab and Ag detection, along with a novel viral 

disruption technique, the most sensitive HIV point-of-care screening 

test will be achieved, at a cost point that enables broad roll-out. 

Note that differences in the ―direct‖ method data between Table 1 and 

Table 2 are the result of an improved capture mAb print buffer instituted 

prior to the later Table 1 experiment. We also note that HIV Ab detection assay does not appear to be 

adversely affected by the detergent treatment (data not shown).

HIV-1/2 Antibody Assay Improvements. Since the goal of Phase II will be integration of an HIV 1/2 Ag/Ab 

combo assay, we include antibody detection results here as well. The 3rd generation MBio rapid HIV-1 Ab 

assay protocol has been simplified by removal of the post-incubation wash step. Performance of the onestep whole blood assay demonstrated using 59 whole blood samples. Samples from 34 HIV-positive donors 

were collected at the UCSD AVRC. Participants provided a venous whole blood tube under an IRBapproved protocol. Tubes were shipped overnight to MBio and assayed the same day as receipt. 25 HIVnegative samples were sourced commercially and were assayed in triplicate. Fig. 5(a) boxplots show Ab 

assay signal intensities for a gp41 antigen sandwich assay for the whole blood samples. Positive and 

negatives are well differentiated. Preliminary work on HIV-2 Ab detection has also been performed, 

focusing on detection of antibodies to the highly conserved env gp36 ectodomain. Although we have just 

recently begun the investigation of gp36 antigens in our 3rd generation serology assay format (and have not 

yet built a significant HIV-2 sample set), early results are promising, as shown in Fig. 5(b). 

MBio HIV 

p24 Ag 

(direct)

MBio HIV 

p24 Ag 

+VDSA

Sample I.D.* S/CO S/CO

177-SL (acute) 0.6 1.9

189-SL (acute) 0.1 1.0

190-SL (acute) 1.7 12.0

191-SL (acute) 2.3 16.0

198-SL (acute) 0.3 0.7

*Acute samples are RNA+ / EIATable 2. MBio p24 Ag assay 

sensitivity comparison. Effect of virus 

disruption sample additive

Research Strategy Page 72

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Figure 5. (a) Whole blood HIV-1 Ab reactivity results for known negative (NEG, N=25) and known positive (POS, 

N=34) samples, reported as signal intensity of gp41 antigen sandwich. (b) HIV-2 Ab reactivity results for known HIV-1 

positive control, HIV-2 positive control, and normal serum, reported as signal of gp36 antigen sandwich. Control 

samples were run in triplicate; error bars are ± one stdev. Different normalization methods result in the absolute signal 

intensity differences between (a) and (b).

PHASE II EXPERIMENTAL RESEARCH DESIGN AND METHODS 

Our goal is to develop and validate a 4th gen HIV-1/2 Ag/Ab combination assay that meets or 

exceeds the performance of clinical laboratory assays, in a low-cost, low-complexity, POC format. 

Strategy. The general strategy for achieving the Phase II project goals is to finalize the selection of 

antibody and antigen reagents, to reduce the complexity of the p24 Ag detection component to the level of 

the demonstrated HIV-1 Ab assay protocol, and to then combine the HIV-1/2 serology and p24 Ag 

immunoassay components on a single POC platform. Improvements in assay sensitivity will be sought 

through a re-design of the cartridge channel and the introduction of a virus disruption reagent. Further 

protocol simplification, to a level that meets FDA CLIA waiver guidelines, will be accomplished through 

cartridge design modifications and the development of a method for on-cartridge lyophilization of assay 

reagents. If necessary to reach performance goals, a mechanism for washing the cartridge waveguide 

array without user intervention will be integrated into the final cartridge and existing reader instrument. The 

integrated system will then be validated with a large carefully chosen set of HIV infection and control 

samples with pre-market evaluation in a clinical setting. 

Specific Aim 1: Assay Development. Combine the Phase I p24 antigen detection assay with the MBio 

multiplexed serology assay cartridge, addressing issues of final monoclonal antibody (mAb) pair 

selection, HIV-1 Ag and HIV-2 Ag selection, reagent conjugations, cross-reactivity, and minimization of 

assay steps and complexity. The Aim 1 milestone is an HIV-1/2 antigen/antibody detection assay with 

performance equivalent to FDA-approved laboratory 4th gen Ag/Ab combo assays for the MBio 

early/acute sample collection.

Selection of p24 antibody pair. Although significant progress has been made on the identification of a high 

affinity anti-p24 monoclonal antibody sandwich pair, the search has not been exhaustive. Additional 

candidate antibodies will be tested as both detection and capture agents. Availability of long term supply 

agreements will be part of the mAb evaluation. Antibody-dye conjugation will be optimized at MBio. A 

design of experiments (DOE) approach will be used to optimize Ab print buffers. Dye labeled recombinant 

p24 and anti-species Ab will be used as fluorescent probes, and spot signal/noise, morphology (intra-spot 

%CV), and lot-to-lot reproducibility as primary metrics. A protocol that mimics the steps of the current HIV-1 

Ab assay will be followed. Briefly, the sample will be combined with detect antibody and added to the 

cartridge entrance port. Following a 20-min incubation period, a wash volume will be added and the 

cartridge will be imaged on the MBio Array Reader. In addition to the NIBSC p24 reference standard, 

additional control samples from the Phase 1 collection will be validated for assay development. p24 antigen 

Known HIV-1 Ab Reactivity

NEG POS

Signal Intensity (arb. units)

-100

0

100

200

300

400

500

0

2

4

6

8

10

12

14

16

18

20

HIV-1 HIV-2 Normal Serum

Signal Intensity (arb. units)

Research Strategy Page 73

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
detection sensitivity and specificity will be analyzed with a three-point titration of the NIBSC p24 standard 

and an 8-member panel of HIV-negative serum comprised of four non-heterophilic antibody (HA) samples 

and four known HA-containing samples. Iterative screening will be performed. 

Results from a titration of the A647-labeled detect Ab will inform a decision on whether substituting the 

significantly brighter SLP3 for A647 will be necessary to meet assay performance goals. If yes, Columbia 

Biosciences (the SLP3 source) has provided MBio with a quote for custom conjugation of SLP3 to a 

supplied antibody. 

Selection of HIV-1 and HIV-2 antigens. The MBio 3rd gen HIV-1 Ab assay utilizes purified recombinant HIV1 gp41 protein from Fitzgerald. While assay performance with this antigen preparation has been 

satisfactory, alternative antigens will be tested in order to further improve performance and to ensure 

multiple reliable sources for product reagents. MBio is currently in a confidential contract negotiation with a 

cGMP supplier of HIV-1 and -2 recombinant proteins. Antigenic peptides will also be considered, as 

improved HIV-1/HIV-2 specificity may be achieved (16, 17). The screening path will be similar to that 

followed for anti-p24 mAb selection, including incorporation of print buffer optimization. Detect antigens will 

be labeled with A647 (or an equivalent dye). The assay sensitivity and specificity provided by the candidate 

HIV-1 and HIV-2 antigens will be measured by assaying an anti-HIV-1/2 combo performance panel 

(Seracare # PRZ207). Non-specific binding of candidate capture antigens to the anti-p24 detect mAb, and 

of the anti-p24 capture mAb to candidate detect antigens, will be investigated in detail. 

Concentration of detect antigens. The stoichiometry of the labeled detect Ag and sample target antibody 

heavily influences assay sensitivity. In low titer samples, a high detect Ag concentration can saturate Ag 

binding sites thus preventing the antibody from serving as a bridge between the immobilized capture Ag and 

the detect Ag. Conversely, with very high Ab titer samples, a relatively low detect Ag concentration can 

result in most captured antibodies being unbound to detect Ag (―hook effect‖). The Phase I HIV-1 Ab assay 

results (Table 1) suggest that a lower detect Ag concentration would have improved assay sensitivity. High 

titer Ab samples yielded s/co values of approximately 36; a two-fold reduction in detect Ag concentration 

would have increased the signals on low titer samples without sacrificing detection of the high titer samples. 

The selected HIV-1 and HIV-2 detect antigens will be carefully titrated with a panel of very high titer and 

very low titer samples to arrive at their final assay concentrations. 

Addition of a virus disruption step. To improve the sensitivity of p24 immunoassays, samples are often 

subjected to detergents and heat, or acid followed by neutralization, to release Ag from both anti-p24 

antibodies and viral particles (18, 19). These antibody-denaturing methods are not, however, available to 

the proposed rapid 4th gen assay. In the preliminary studies section, a detergent-based virus disruption 

sample additive (VDSA) was described that improved p24 Ag detection with a small set of acute infection 

samples without adversely affecting the serology component of the assay. An Aim 1 activity will be 

development of the VDSA reagent for best performance, including development of lyophilized, heat stable 

formulations. Reagent lyophilization is discussed in more detail under Aim 2. 

Combining HIV-1/2 Ab detection and p24 Ag detection. The current anti-gp41 Ab and p24 Ag detection 

assays share a common sample additive buffer (minus HA-blockers for the serology assay) and wash 

buffer. Post-print processing of waveguides and cartridge assembly is identical for the two assay formats. 

Challenges with combining the detection reagents within a single array and sample additive should 

therefore be limited to cross-reactivity issues. While cross-reactivity will be addressed in the final antibody 

and antigen selections, adjustments in the sample additive buffer and wash buffer formulations may be 

required. Incubation times are also similar between the existing assays and it is expected that a mutually 

effective incubation time of between 10 to 20 min will be targeted. 

The envisioned minimal assay protocol will consist of: (1) addition of sample to cartridge entrance port; (2) 

following an incubation period (10-20 min), insert the cartridge into fluorescence reader and, 15-30 sec 

later, receive assay results. A distinguishing advantage of the planar waveguide technology is only 

fluorescent molecules on, or very near, the waveguide service is exposed to the input light; thus free fluor in 

Research Strategy Page 74

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
the solution above the array is not significantly excited. The whole blood assay results discussed in the 

preliminary studies section were obtained with a version of this simple protocol. However, an assay 

performance boost can be obtained with a single wash of the array surface. Whether this post-incubation 

wash step is necessary to meet assay performance goals will be determined under Specific Aim 1. 

Assay performance. The FDA-approved Bio-Rad GS HIV Combo Ag/Ab EIA run at MBio will serve as the 

reference test for Aim 1 method development. Performance of the MBio HIV-1/2 Ag/Ab combination assay 

will be tested with a set of 5 commercially available HIV-1 seroconversion/performance panels (SeraCare 

PRB109 and PRB965; ZeptoMetrix 9075, 9079, and 12008), two anti-HIV-1/2 combo performance panels 

(SeraCare PRZ207 and PRZ208), and an anti-HIV-2 performance panel (SeraCare PRF203). We have also 

secured an expanded, 80 sample ―challenge panel‖ from subaward collaborator Dr. Susan Little at UCSD. 

Samples were selected from a large set (2,755 samples) collected during a prospective HIV screening 

program (NIH P01-AI074621; The San Diego Early Test program (20)). The 80 samples are HIV rapid-test 

negative, and have been characterized by HIV NAT (PCR) and Abbott ARCHITECT HIV Ag/Ab Combo. 

Expected outcomes, risks, and alternatives. The expected outcome of Specific Aim 1 is the 

demonstrated performance (clinical sensitivity and specificity) of a rapid HIV 1/2 Ag/Ab combo assay that is 

equivalent to that of an FDA-approved laboratory-based 4th gen assays. By reducing the complexity of the 

p24 Ag detection component of the assay, we risk losing the assay sensitivity required for early Ag 

detection in acute infection samples. It is expected, however, that direct conjugation of SLP3 to the detect 

Ab, a re-design of the cartridge channel (Aim 2), and incorporation of an VDSA reagent will more than 

recover any lost assay sensitivity. If an increase in sensitivity is still required, the potential of utilizing 

biotinylated detect reagents paired with delivery of SA-SLP3 from an on-cartridge reagent reservoir will be 

investigated. 

Specific Aim 2: Cartridge Integration. Modify the MBio Cartridge, Rack, Reader, and Software to deliver 

an automated HIV-1/2 Ag/Ab combo result, and incorporate heat stable assay reagents into the MQ 

cartridge. The Aim 2 milestone is a portable, integrated system delivered to clinical collaborators that 

meets FDA CLIA waiver guidance requirements. 

Representative MBio cartridge components are shown here. Location of lyophilized 

reagents is not shown (confidential). A detailed description of each component is 

beyond scope this proposal. Modifications to the assay cartridge will address liquid 

flow, sample loading, introduction of test quality controls, on-cartridge lyophilized 

assay reagents, and potentially integration of a wash buffer reservoir. 

Fluidic channel geometry for improved assay sensitivity. The current cartridge 

geometry places 0.6 mm diameter printed antibody or antigen features within a fluid 

channel with a width of 3 mm and height of 0.145 mm. The time between sample 

addition and channel filling via capillary flow is only 3-4 seconds. On-rate and 

diffusion limitations to analyte binding should be reduced by both reducing the 

volume of the channel and the flow rate. With existing cartridge components, 

channel geometries can be altered by simply changing the dimensions of a die-cut 

adhesive gasket. The assay performance effect of 2 mm width channels with heights 

of 0.145, 0.1, 0.075 and 0.50 mm will be tested. The reduced channel volumes will 

both slow flow rate and ―funnel‖ a greater fraction of the sample over printed proteins. 

Sample Loading. Robustness of sample loading is a critical design feature in a CLIA-waivable test. The 

cartridge will be design to directly accept finger stick whole blood, without the use of transfer pipets. The 

inlet port will be designed such that direct contact with the blood droplet will result in capillary fill into the 

cartridge. The same inlet port will be designed to be compatible with a range of transfer devices (e.g., 

pipets) when the sample is a blood tube. The approximate blood volume from a finger stick sample is 17 l 

(21). With the current 3 mm wide X 0.145 mm high cartridge channel, an 18-l sample volume is sufficient 

to cover the planar waveguide array. The channel re-design described under Aim 1 should reduce this 

Research Strategy Page 75

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
required volume to not more than 10 l. The design goal is to have the assay perform with precision for a 

sample volume range of 10 to 30 l. 

Internal Quality Controls. On-cartridge controls will be integrated to provide confirmation of proper sample 

introduction and reagent formulation. The addition of anti-detect reagent antibodies to the printed array, 

downstream of the array capture reagents, will serve as reagent formulation controls (e.g., anti-mouse IgG 

Ab, anti-gp41Ab and anti-gp36 Ab). 

On-Board Reagents. On-cartridge dried reagents will be necessary to meet FDA CLIA waiver guidance 

requirements and to achieve long-term storage and thermal stability. Formulations and lyophilization 

conditions are separately being developed at MBio to obtain stable products, and this will be adapted for the 

proposed device. A requirement is that the dried reagents rapidly hydrate and disperse evenly throughout 

the sample, properties that will be targeted with a careful study of excipients and caking agents. An 

additional requirement is that the sample volume brought in contact with lyophilized reagent is controlled. 

MBio cartridges use controlled hydrodynamic forces (channel geometry and tuned surface energy) and 

capillary gates to control flow. For the current whole blood cartridge, the channel fills and then flow stops 

once the end of the channel is reached. The head pressure of excess sample volume in the entrance port 

is not sufficient to push sample volume into the exit port, even when the cartridge is held vertically. 

Dried reagents will be maintained by sealing cartridges in vapor barrier pouches with desiccant material. 

MBio has the necessary temperature- and humidity-controlled environmental chambers for testing of both 

real time and accelerated stability. If the assay protocol demands a wash step, a means to do so without a 

manual intervention step will be developed. One approach is to place a ―blister pack‖ containing the wash 

buffer on the cartridge. The blister pack will sit above a reservoir and channel that directs the flow of wash 

buffer to the channel. A simple solenoid-driven plunger in the instrument would depress the blister pack 

releasing the buffer contents. 

Expected outcomes, risks, and alternatives. Aim 2 activities are expected to produce an assay system 

that delivers HIV-1/2 Ag/Ab combo assay results with only two user intervention steps- addition of sample to 

the assay cartridge and insertion of the cartridge into the reader instrument. The main risk is that 

reconstitution of the lyophilized reagents is not sufficiently rapid and/or does not yield a reproducible 

concentration of these reagents in the cartridge channel volume. Alternative cartridge designs that further 

restrict sample volume flow as well as alternative caking reagents and lyophilization protocols would be 

investigated as correcting measures. An additional risk is that a post-incubation wash becomes a 

requirement and that the delivery of wash buffer from a blister pack is unsuccessful. An alternative 

approach would be the addition of wash buffer to the entrance port with a dropper bottle. Although this 

would require a manual intervention step, the volume would not require metering. 

Specific Aim 3: Assay Validation. Validate system using well characterized early HIV infection specimens 

including a panel of 200 HIV positive specimens comprised of 20 acute infection samples (RNA+ / Ab -), 

early seroconversion (Western Blot indeterminate) and seropositive (HIV-1 and HIV-2) samples. 200 

HIV-negative samples will be used for specificity testing. The Aim 3 milestone is a dataset 

demonstrating performance equivalent to FDA-approved laboratory 4th gen HIV-1/2 Ag/Ab assays. 

The performance of the developed assay system will be validated by assaying a panel of ≥ 200 HIV-positive 

samples, including a minimum of 20 acute infection samples (HIV RNA+ / Ab -) sourced through 

collaborator at UCSD. The remainder of the set will include early infection samples, high Ab titer samples, 

commercially available HIV-1 subtype performance panels, a commercially available HIV-1 

Incidence/Prevalence panel, and the HIV-1 and HIV-2 panels listed under Aim 1. 200 HIV-negative (RNA-) 

samples will be acquired from Valley BioMedical and Dr. Susan Little for specificity testing. Assume 95% 

sensitivity and specificity, these sample numbers give 95% confidence intervals of ± 3%. In addition, it will 

be determined whether a panel of the following potentially interfering substances have effects on assay 

performance: lipemic samples (≥ 1000 mg/dL triglycerides, icteric samples (≥ 20 mg/dL bilirubin), 

hemolyzed samples (≥ 500 mg/dL hemoglobin), and proteinemic samples (≥ 12 g/dL protein). 

Research Strategy Page 76

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Expected outcomes, risks, and alternative. The objective of Aim 3 is to demonstrate that the 

performance (clinical sensitivity and specificity) of the rapid HIV 1/2 Ag/Ab combo assay system is that is 

equivalent to FDA-approved 4th gen assays (95% confidence intervals match the Bio-Rad GS HIV-1/2 

Ag/Ab Combo. The main risk is that performance does not meet expectations and that additional system 

development is required. In the event that performance equivalent to 4th gen laboratory tests is not 

achieved, system advantages in the context of a 3rd gen HIV assay relative to FDA-approved rapid tests will 

be pursued; 3rd gen product launch is considered low technical risk given existing data (e.g. Table 1). 

Specific Aim 4 - Pre-Market Evaluation. Place systems in intended use setting and capture operational 

and usability feedback in advance of design lock; and generate a preliminary dataset on capillary whole 

blood samples from 100 study participants in San Diego. The Aim 3 and 4 milestone is a system design 

and dataset for an FDA investigational device exemption (IDE) meeting in advance of clinical trials. 

The Pre-Market Evaluation will take place at UCSD HIV care clinics (AVRC and Owen Clinic), under the 

direction of subaward co-investigators Benson and Little, who will direct AVRC staff. A study of 

approximately 100 finger stick donors will be designed. The study will accomplish two goals: (1) generate 

usability dataset, and (2) establish performance in intended use setting. 

Usability Testing. Given that intended use is a CLIA-waived product that will be operated by nonprofessional staff, a first aspect of the pre-market evaluation will be an assessment of how operators 

interact with the system, including patient, cartridge, and reader user interface/software. Human factors 

analysis will follow FDA and ISO 13485 guidance, and will be based on surveys and direct observation. 

Usability testing will be performed in the context of MBio‘s Quality Management System, and will be part of 

the Design and Risk Management Processes. 

Intended Use Setting Performance. Although not a powered clinical study, the analysis of 100 study 

participants will provide critical feedback on system performance. Donors will provide both finger stick and 

venous samples. (NOTE: Human Subjects topics are addressed in a separate section of this proposal). 

Sensitivity relative to the FDA approved rapid tests and the Bio-Rad GS HIV-1/2 Ag/Ab Combo will be 

established. Reproducibility of the system will be established by running venous samples in triplicate. 

Importantly, accuracy and precision of finger stick samples relative to venous will be established. 

The outcome of Phase II should be a system design and dataset that the company will use as the basis for 

a product launch decision. Data generated under Phase II will form the basis of the pack MBio takes to a 

preliminary FDA investigational device exemption (pre-IDE) meeting in advance of clinical trials. 

PROGRAM TIMELINE Yr1: Q1 Yr1: Q2 Yr1: Q3 Yr1: Q4 Yr2: Q1 Yr2: Q2 Yr2: Q3 Yr2: Q4

Aim 1: Assay Development

1.1 p24 mAb selection / optimization

1.2 HIV-1 and -2 antigen selection

1.3 Viral disruption assay

Aim 2: Cartridge Integration

2.1 Fluidic channel geometry

2.2 Sample loading / inlet port design

2.3 Internal QC

2.4 On-board reagents

Aim 3: Assay Validation

3.1 HIV positive panel

3.2 HIV negative panel

3.3 Interfering substance

Aim 4: Pre-Market Field Evaluation

4.1 Study plan

4.2 Usability testing

4.3 Prospective study (100 donors)

Research Strategy Page 77

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Human Subjects 

All studies involving human subjects conducted under this project will comply with federally-mandated human 

subjects regulations as presented in 45 CFR 46 (Regulations for Research Involving Human Subjects). In 

addition, the project will also be in compliance with OHRP guidance regarding the use of repositories for 

human serum, tissue, urine and other body fluids. All clinical sites that recruit subjects for donation of 

specimens will hold a Federal Wide Assurance (FWA) and strictly enforce the protections outlined for human 

subjects in accordance with the FWA and local policy. Key personnel who participate in clinical research are 

required to complete education in protection of human research participants which is recognized by the NIH 

and acceptable to local human subjects review boards. Certification of human subjects training will be kept on 

file for all key personnel and will be provided upon request. No research will be implemented without prior 

approval of the local Institutional Review Board or Ethics Committee or other relevant regulatory. 

Risks to Subjects 

Involvement and Characteristics of Human Subjects. Patients recruited to donate specimens and for whom 

clinical data will be collected for assay studies will include male and female adults (> 18 years of age) who are 

at risk for, infected with, or being tested for human immunodeficiency virus (HIV) and willing and able to 

provide informed consent. The health status of study participants from whom specimens have been or will be 

obtained will include those with signs or symptoms of acute HIV infection and asymptomatic or otherwise 

healthy individuals. Specific protocols will note the eligibility, inclusion, and exclusion criteria for recruitment to 

the project. Vulnerable populations such as prisoners and children will not be included as they will derive no 

discernable benefit from study participation, and will not be able to provide independent informed consent. 

Pregnant women, if they are undergoing testing for HIV and are willing and able to provide informed consent, 

may be referred to the project and will be offered the opportunity to donate specimens for this project. 

Pregnancy is unlikely to alter performance characteristics of the assay development, and in this context 

pregnant women will be eligible to participate because there is reasonable expectation that the assay being 

evaluated will have no adverse consequences to the woman or her fetus. 

Sources of Research Material. Likely sources of research material will include samples of blood, serum, or 

other sterile body fluids obtained from study volunteers or archived in the UCSD Acute and Early HIV infection 

(AEH) Program repository with patient consent for future testing on file. Patients’ existing medical records will 

be reviewed to obtain correlative clinical data. Surveys or questionnaires may also be used as a source of 

research data. The clinical data collected will be used solely for the purpose of research as defined by the 

project. Most of this information, however, will be available to the clinicians providing care to the patients if 

needed. All data, including laboratory data, will be identified by a patient-specific patient identification number 

(PID). Data submitted to the project database will not bear patient identifiers. Only local study personnel will 

have access to any link between PID and patient identifiers. Aliquoted specimens may be retained at the study 

site for use in research studies that are necessary to support the primary findings of the studies, and any future 

research studies of stored specimens will be allowed only after specific proposals for their use are reviewed 

and approved by the project, its partners and the NIH project officer have determined that use is covered by 

existing informed consent. 

Potential Risks of Study Participation. Possible physical risks to study participants are limited to those 

associated with venipuncture for blood samples, interviews, and questionnaires. The risk from these 

procedures is considered minimal. It is possible that psychological trauma may result from a diagnosis of HIV 

infection. Potential social risks include embarrassment and stigmatization if there is inadvertent disclosure of 

confidential information such as diagnosis of HIV. The informed consent for obtaining samples and data for this 

project will include the alternatives to study participation and these will be reviewed with potential study 

subjects, as will risks and benefits. 

Adequacy of Protection Against Risks 

Recruitment and Informed Consent. Recruitment strategies for the project will be determined by the clinical 

investigators and recruitment will occur only after full approval for the study is obtained from the local 

Institutional Review Board or Ethics Committee and other relevant regulatory entities. Written informed consent 

documents for participation provide complete descriptive information about the research. They will be 

translated in a language that the participant understands as needed, and will be back translated into English to 

assure the adequacy of the translation. If potential study participants are unable to read, the clinical study site 

Protection of Human Subjects Page 78

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
will comply with their local Institutional Review Board or Ethics Committee guidelines for obtaining informed 

consent. In addition to the written document, verbal discussions will take place between the study participant 

and investigators before the consent form is signed. Additional information to be provided in the informed 

consent document includes adverse effects, risks, painful procedures, and the need for any follow-up visits. 

Compensation will also be addressed in the consent form. After approval of the consent form, written informed 

consent will be obtained by a member of the medical research team from the study participant. The signed 

informed consent document will be maintained in the patient's medical record and the study site source 

document; the patient will receive a copy. The informed consent process will be documented in each 

participant’s study chart, and where appropriate, additional IRB-approved educational or recruitment materials 

will be provided to the participants. All clinical care required for study participants identified with HIV who agree 

to donate samples and clinical data to this project will be at the discretion of their primary physicians. Any 

relevant information necessary to their clinical care will be provided to the primary physician in a timely 

manner. 

Protections Against Risk. Efforts will be made to limit any psychological impact for the study participant and his 

or her family. The investigators will maintain all patient information in strict confidence. All information about 

study participants will be contained in the medical record that is maintained in a secure facility. All research 

records will identify the patient only by a coded number. The master linkage between patient name and coded 

number will be accessible only by study personnel. We will follow all guidelines for confidentiality in research. 

Potential Benefits of the Proposed Research to the Subjects and Others 

Benefits of participation may include the provision of information that could result in more rapid identification 

and diagnosis of HIV that might afford appropriate referral for treatment or other interventions to reduce 

morbidity. The proposed research may result in the development of new technology that may allow earlier 

detection of HIV, which might benefit public health by reducing risk of transmission of HIV from infected 

persons to others. 

Importance of Knowledge to be Gained 

The proposed research may result in the development of new technology that might allow earlier detection of 

HIV, which might benefit public health by reducing transmission of HIV from infected persons to others and 

personal health by allowing earlier initiation of appropriate treatment for HIV. 

Data and Safety Monitoring Plan 

Data and safety will be monitored on multiple levels. At the individual institution, we will monitor patients for the 

occurrence of any adverse events associated with specimen or data collection and report these events as 

directed by the institution’s adverse event reporting policy. Any event is reported directly to the study team. 

Data collection and data integrity will be monitored by the team through conference calls and review of data at 

face to face meetings. NIH project officers will be included in the data monitoring plans. 

Inclusion of Women and Minorities in Research 

Women and minorities will be appropriately represented in patient populations recruited to provide specimens 

for project studies. The substantial portion of our research participants are likely to be racial/ethnic minorities, 

based on HIV demographics in our region. The populations will be typical of the HIV epidemic in San Diego 

County, and in each additional area from which specimens are recruited. There will be representation of both 

males and females in project studies, and the distribution by sex will reflect nearly the proportions affected or at 

risk for HIV based on the demographics of local statistics. There are no data currently to suggest that 

diagnostic test results will differ by sex. Any differences will be explored in secondary analyses of study results. 

In addition, we will also conduct exploratory analyses to determine any racial/ethnic differences where the 

enrolled population allows. 

Inclusion of Children in Research 

Children will not be included in the research, based on their general inability to provide independent informed 

consent and provision of high quality samples for use in the assays. Should a sensitive and specific assay be 

produced as a result of this research, children will be included in the next phases of diagnostic testing. 

Protection of Human Subjects Page 79

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Inclusion of Women and Minorities in Research 

Women and minorities will be appropriately represented in patient populations recruited to provide specimens 

for project studies. The substantial portion of our research participants are likely to be racial/ethnic minorities, 

based on HIV demographics in our region. The populations will be typical of the HIV epidemic in San Diego 

County, and in each additional area from which specimens are recruited. There will be representation of both 

males and females in project studies, and the distribution by sex will reflect nearly the proportions affected or at 

risk for HIV based on the demographics of local statistics. There are no data currently to suggest that 

diagnostic test results will differ by sex. Any differences will be explored in secondary analyses of study results. 

In addition, we will also conduct exploratory analyses to determine any racial/ethnic differences where the 

enrolled population allows. 

Women &Minorities Page 80

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Targeted/Planned Enrollment Table

This report format should NOT be used for data collection from study participants.

Study Title: HIV Antigen/Antibody Diagnostic

Total Planned Enrollment: 100

TARGETED/PLANNED ENROLLMENT: Number of Subjects

Ethnic Category Sex/Gender

Females Males Total

Hispanic or Latino 4 12 16

Not Hispanic or Latino 6 78 84

Ethnic Category: Total of All Subjects * 10 90 100

Racial Categories

American Indian/Alaska Native 0 2 2

Asian 0 4 4

Native Hawaiian or Other Pacific Islander 2 4 6

Black or African American 4 8 12

White 6 70 76

Racial Categories: Total of All Subjects * 12 88 100

* The “Ethnic Category: Total of All Subjects” must be equal to the “Racial Categories: Total of All Subjects.”

Planned Enrollment Table Page 81

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
Inclusion of Children in Research 

Children will not be included in the research, based on their general inability to provide independent informed 

consent and provision of high quality samples for use in the assays. Should a sensitive and specific assay be 

produced as a result of this research, children will be included in the next phases of diagnostic testing. 

Children Page 82

Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
BIBLIOGRAPHY 

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transmission per coital act, by stage of HIV-1 infection, in Rakai, Uganda. J Infect Dis. 

2005;191(9):1403-9.

2. Fiebig EW, Wright DJ, Rawal BD, Garrett PE, Schumacher RT, Peddada L, et al. Dynamics of HIV 

viremia and antibody seroconversion in plasma donors: implications for diagnosis and staging of 

primary HIV infection. AIDS. 2003;17(13):1871-9.

3. CLSI. Criteria for Laboratory Testing and Diagnosis of Human Immunodeficiency Virus Infection: 

Approved Guideline. CLSI. 2011;M53-A.

4. Patel P, Bennett B, Sullivan T, Parker MM, Heffelfinger JD, Sullivan PS. Rapid HIV screening: 

missed opportunities for HIV diagnosis and prevention. J Clin Virol. 2012;54(1):42-7.

5. CDC. HIV / AIDS in the United States. 2012. 

6. WHO. Global Summary of the AIDS Epidemic; World Health Organization and UNAIDS. 2007. 

7. Louie B, Lei J, Liska S, Dowling T, Pandori MW. Assessment of sensitivity and specificity of first, 

second, and third generation EIA for the detection of antibodies to HIV-1 in oral fluid. J Virol 

Methods. 2009;159(1):119-21.

8. Pandori MW, Hackett J, Jr., Louie B, Vallari A, Dowling T, Liska S, et al. Assessment of the ability of 

a fourth-generation immunoassay for human immunodeficiency virus (HIV) antibody and p24 antigen 

to detect both acute and recent HIV infections in a high-risk setting. J Clin Microbiol. 

2009;47(8):2639-42.

9. Pandori MW, Louie B, Pilcher C, Klausner JD, Vallari A, Harris B, et al. Assessing the sensitivities of 

laboratory-based and point-of-care HIV antigen-antibody combination tests using a panel of 

specimens from recently and acutely infected individuals. 2010 HIV Diagnostics Conference; 

Orlando, FL2010. 

10. Kilembe W, Keeling M, Karita E, Lakhi S, Chetty P, Price MA, et al. Failure of a novel, rapid antigen 

and antibody combination test to detect antigen-positive HIV infection in African adults with early HIV 

infection. PLoS One. 2012;7(6):e37154. PMCID: 3371037. 

11. Chetty V, Moodley D, Chuturgoon A. Evaluation of a 4th generation rapid HIV test for earlier and 

reliable detection of HIV infection in pregnancy. J Clin Virol. 2012;54(2):180-4.

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infection: a field evaluation of the determine(R) HIV-1/2 Ag/Ab combo test. J Infect Dis. 

2012;205(4):528-34. PMCID: 3318673. 

13. Taegtmeyer M, MacPherson P, Jones K, Hopkins M, Moorcroft J, Lalloo DG, et al. Programmatic 

evaluation of a combined antigen and antibody test for rapid HIV diagnosis in a community and 

sexual health clinic screening programme. PLoS One. 2011;6(11):e28019. PMCID: 3222669. 

14. Tang S, Hewlett I. Nanoparticle-based immunoassays for sensitive and early detection of HIV-1 

capsid (p24) antigen. J Infect Dis.201 Suppl 1:S59-64.

15. Nabatiyan A, Baumann MA, Parpia Z, Kelso D. A lateral flow-based ultra-sensitive p24 HIV assay 

utilizing fluorescent microparticles. J Acquir Immune Defic Syndr. 2010;53(1):55-61.

16. Workman S, Wells SK, Pau CP, Owen SM, Dong XF, LaBorde R, et al. Rapid detection of HIV-1 p24 

antigen using magnetic immuno-chromatography (MICT). J Virol Methods. 2009;160(1-2):14-21.

17. Miedouge M, Greze M, Bailly A, Izopet J. Analytical sensitivity of four HIV combined 

antigen/antibody assays using the p24 WHO standard. J Clin Virol. 2011;50(1):57-60.

18. Lochhead MJ, Todorof K, Delaney M, Ives JT, Greef C, Moll K, et al. Rapid Multiplexed 

Immunoassay for Simultaneous Serodiagnosis of HIV-1 and Co-Infections. J Clin Microbiol. 

2011;49(10):3584–90.

19. Fenouillet E, Sorensen AM, Lacroix M, Coutellier A, Herson S, Fretz-Foucault C, et al. Early and 

specific diagnosis of seropositivity to HIVs by an enzyme-linked immunosorbent assay using envderived synthetic peptides. AIDS. 1990;4(11):1137-40.

20. Petrov RV, Khaitov RM, Sidorovich IG, Pavlikov SP, Nikolaeva IA, Ivachenko ME, et al. The use of 

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21. Schupbach J, Flepp M, Pontelli D, Tomasik Z, Luthy R, Boni J. Heat-mediated immune complex 

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detection in plasma as sensitive as HIV-1 RNA detection by polymerase chain reaction. AIDS. 

1996;10(10):1085-90.

22. Schupbach J, Tomasik Z, Knuchel M, Opravil M, Gunthard HF, Nadal D, et al. Optimized virus 

disruption improves detection of HIV-1 p24 in particles and uncovers a p24 reactivity in patients with 

undetectable HIV-1 RNA under long-term HAART. J Med Virol. 2006;78(8):1003-10.

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antibodies, antigens, and nucleic acids for diagnosis of acute HIV infection. J Clin Microbiol. 

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24. Peck HR, Timko DM, Landmark JD, Stickle DF. A survey of apparent blood volumes and sample 

geometries among filter paper bloodspot samples submitted for lead screening. Clin Chim Acta. 

2009;400(1-2):103-6.

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Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J
PHS 398 Checklist

OMB Number: 0925-0001 

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the questions that are specific to the PHS398.

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4. * Program Income

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Principal Investigator/Program Director (Last, first, middle): Lochhead, Michael, J