Rapid, Onsite COVID-19 Detection
Phase: NA
Conditions: COVID-19, Sars-CoV2
Sponsor: University of Wisconsin, Madison



[Rapid, onsite COVID-19 detection] 
 
[O’Connor] 
 
 
 
Version #: [3] 
Version Date: [08/30/2020] 
Page 1 of 1 
 
1.0 
STATEMENT OF COMPLIANCE 
I confirm that I have read this protocol. I will comply with the IRB-approved protocol, and applicable 
regulations, guidelines, laws, and institutional policies.  
 
I agree to ensure that all staff members involved in the conduct of this study are informed about their 
obligations in meeting the above commitment.  
 
Name 
Signature 
Date 
 
 
__________________________________ 
_____________________________ 
___________________ 
Principal investigator 
 
 
__________________________________ 
_____________________________ 
___________________ 
Sponsor 
 
 
 
 
 
 

[Rapid, onsite COVID-19 detection] 
 
[O’Connor] 
 
 
 
Version #: [3] 
Version Date: [08/30/2020] 
Page 1 of 1 
 
2.0 
LIST OF ABBREVIATIONS 
AE 
Adverse Event 
AVRL 
AIDS Vaccine Research Laboratory 
BSC 
Biological Safety Cabinet 
CCC 
Clinical Coordinating Center 
CFR 
Code of Federal Regulations 
CRF 
Case Report Form 
CTMS 
Clinical Trial Management Software 
DCC 
Data Coordinating Center 
DHHS 
Department of Health and Human Services 
DMC 
Data Monitoring Committee 
EtOH 
Ethyl alcohol 
FDA 
Food and Drug Administration 
GCP 
Good Clinical Practice 
HIPAA 
Health Insurance Portability and Accountability Act 
IDE 
Investigational Device Exemption 
ICTR 
Institute for Clinical and Translational Research 
IRB 
Institutional Review Board 
NIH 
National Institutes of Health 
OHRP 
Office for Human Research Protections 
PHI 
Protected Health Information 
PI 
Principal Investigator 
POC 
Point of Contact 
RT-LAMP 
Reverse transcription loop-mediated isothermal amplification 
SAE 
Serious Adverse Event 
SAP 
Statistical Analysis Plan 
SMC 
Safety Monitoring Committee 
SMP 
Study Monitoring Plan 
SMS 
Study Monitoring Service 
UP 
Unanticipated Problem 
UWCCC 
University of Wisconsin Carbone Cancer Center 
 

[Rapid, onsite COVID-19 detection] 
 
[O’Connor] 
 
 
 
Version #: [3] 
Version Date: [08/30/2020] 
Page 2 of 20 
 
Study Population 
Anyone over the age of 5yrs old with consent to provide a saliva sample will be eligible to 
participate.  
Estimated 
Subject Duration 
The duration of the study for each subject is approximately 20 minutes each visit. 
Estimated 
Enrollment 
Period & Study 
Duration 
Study enrollment will occur over a 12 month period.  
 
 
 

[Rapid, onsite COVID-19 detection] 
 
[O’Connor] 
 
 
 
Version #: [3] 
Version Date: [08/30/2020] 
Page 13 of 20 
 
a. 
Beyond this point, any potential SARS-CoV-2 particles in the saliva can be considered 
inactivated and could be handled without the plexiglass shield or outside of a BSC. 
 
4) While saliva samples are being heat inactivated, prepare the appropriate volume of the following 
Master Mix for the colorimetric RT-LAMP reactions: 
 
Reagent    
 
  
1X Rxn     
9X Rxn     
25X Rxn     
100X Rxn 
____________________________________________________________________ 
WarmStart Colorimetric     
LAMP 2X Master Mix     
(NEB, M1800L)    
  
10 ul      
90 ul      
250 ul    
 1000 ul 
dH2O    
 
         
  5 ul      
45 ul      
125 ul    
   500 ul 
10X LAMP primer 
mix (ColorLAMP N)      
  2 ul      
18 ul       
  50 ul    
   200 ul 
____________________________________________________________________ 
 
5) Aliquot 17 ul of colorimetric RT-LAMP reaction mix into a fresh set(s) of PCR strip-cap tubes. Prepare 
samples in duplicate for each saliva sample. Hold these LAMP reaction tubes in a cooler prior to adding 
heat-inactivated saliva samples. 
 
6) Add 3ul of heat-inactivated saliva (that is already diluted 1:1 in 1X PB) or add 1.5 ul of heat-inactivated 
saliva and 1.5ul of 1x PBS to the LAMP reaction. The final four positions in each batch of LAMP 
reaction strip-cap tubes should be reserved for two negative and two positive controls, respectively.   
- 
The negative control reactions should contain 3 ul water/PBS. 
 
- 
The positive control reactions should include 1000 copies of SARS-CoV-2 virus that has 
been inactivated by heat or UV irradiation. The vRNA is prepared in a 50:50 mix of water 
and 1X PBS to a concentration of 333 copies/ul so that we can add 3 ul to the positive 
control. With 1000 copies of vRNA in the positive control, this would equate to a saliva 
sample concentration of 666,666 copies of SARS-CoV-2 per ml of saliva. Thus, our assay 
will readily detect saliva containing a concentration of 10^6 copies of SARS-CoV-2 per ml.  
 
7) Take a picture of the strip tubes before incubating the RT-LAMP reaction. 
 
8) Incubate the colorimetric RT-LAMP reactions in a thermomixer or dry bath at 65℃ for 30 minutes while 
shaking if needed. 
  
9) Document any color changes at a 30-minute timepoint by digital photography of the RT-LAMP reactions 
against a prelabeled template on a white background. Color change from pink to yellow in both 
replicates is indicative of a sample with SARS-CoV-2 RNA present. If the color change to yellow is only 
clear in one of two replicate saliva samples, the sample will be scored as a potential finding of clinical 
significance. 
 
10) Once a session of RT-LAMP testing is completed, disinfect all lab equipment with 10% bleach wipes 
followed by 70% EtOH spray. Collect all used pipet tips, plasticware, wipes, PPE, etc. in a biohazard