Indocyanine Green Fluorescence Guided Imaging in Lower Extremity Amputation Patients
Conditions: Trauma Injury
Sponsor: Dartmouth-Hitchcock Medical Center

PROTOCOL TITLE: ICG Fluorescence Imaging in Lower Extremity Amputation Patients
D-HH IRB OVERSIGHT:
One of the following must be true in order to submit to the D-HH IRB. Please check all that 
apply:
☒ The Principal Investigator is employed by D-H
☒ The study will utilize any D-H data or specimens
☒ The study will enroll D-H patients or recruit from D-H sites
☒ The study will utilize any D-H resources, e.g. study procedures will occur at 
     D-H locations and/or use of D-H equipment or shared resources
PROTOCOL TITLE:
ICG Fluorescence Imaging in Lower Extremity Amputation Patients
PRINCIPAL INVESTIGATOR:
Ida Leah Gitajn
Department of Orthopaedics
603-650-7484
Ida.Leah.Gitajn@hitchcock.org
VERSION NUMBER/DATE:
Version 5, August 18, 2022
REVISION HISTORY
Revision 
#
Version Date
Summary of Changes
Consent 
Change?
4
12/11/2020
Included consenting of Spanish Speaking 
Subject
Changes make to recruitment, vulnerable 
populations
Requesting a partial waiver of HIPAA for 
recruitment purposes
Yes
5
08/18/2022
Adding SPY PHI and drug receipt change
Yes

PROTOCOL TITLE: ICG Fluorescence Imaging in Lower Extremity Amputation Patients
Table of Contents
1.0
Study Summary ........................................................................................................4
2.0
Objectives* ...............................................................................................................5
3.0
Background*.............................................................................................................5
4.0
Study Endpoints*......................................................................................................5
5.0
Study Intervention/Investigational Agent.................................................................5
6.0
Procedures Involved* ...............................................................................................7
7.0
Data and Specimen Banking*...................................................................................8
8.0
Sharing of Results with Subjects* ............................................................................9
9.0
Study Timelines*......................................................................................................9
10.0
Inclusion and Exclusion Criteria* ............................................................................9
11.0
Vulnerable Populations*...........................................................................................9
12.0
Local Number of Subjects ......................................................................................10
13.0
Recruitment Methods..............................................................................................10
14.0
Withdrawal of Subjects*.........................................................................................10
15.0
Risks to Subjects*...................................................................................................11
16.0
Potential Benefits to Subjects* ...............................................................................11
17.0
Data Management* and Confidentiality.................................................................11
18.0
Provisions to Monitor the Data to Ensure the Safety of Subjects*.........................12
19.0
Provisions to Protect the Privacy Interests of Subjects ..........................................13
20.0
Compensation for Research-Related Injury............................................................14
21.0
Economic Burden to Subjects.................................................................................14
22.0
Consent Process......................................................................................................14
23.0
Process to Document Consent in Writing...............................................................17
24.0
Setting.....................................................................................................................17
25.0
Resources Available ...............................................................................................18
26.0
Multi-Site Research* ..............................................................................................18

PROTOCOL TITLE: ICG Fluorescence Imaging in Lower Extremity Amputation Patients
1.0
Study Summary
Study Title
ICG fluoresce imaging in lower extremity amputation 
patients
Study Design
Prospective nonrandomized trial
Primary Objective
Evaluate changes to bone perfusion in association with 
osteotomy and soft tissue stripping in patients undergoing 
lower extremity amputation
Secondary 
Objective(s)
None
Research 
Intervention(s)/ 
Investigational 
Agent(s)
ICG contrast infusion and ICG-based fluorescence imaging
IND/IDE #
NA
Study Population
Patients undergoing lower extremity amputation
Sample Size
30 patients
Study Duration for 
individual 
participants
During surgical procedure only
Study Specific 
Abbreviations/ 
Definitions
AE: adverse event/adverse experience 
BWH: Brigham and Women’s Hospital 
CT: computed tomography
CRF: case report forms
DCE-FI: dynamic contrast enhanced fluorescence imaging 
DHMC: Dartmouth-Hitchcock Medical Center
GDP: Good Clinical Practice 
ICG: indocyanine green
Imx: maximum ICG fluorescence intensity 
LAR: legally authorized representative
SAE: serious adverse event/serious adverse experience 
SOP: standard operating procedure

PROTOCOL TITLE: ICG Fluorescence Imaging in Lower Extremity Amputation Patients
2.0
Objectives*
Specific Aim 1: Identify bone perfusion kinetics at baseline and following osteotomy and 
subsequent soft tissue stripping in patients undergoing scheduled lower extremity 
amputation.
Hypothesis 1—Human bone perfusion kinetics will demonstrate predictable and 
reproducible changes when undergoing vascular compromise, osteotomy, and soft tissue 
stripping.
Specific Aim 2: Optimize image data processing and identify ICG-based DCE-FI kinetic 
curve properties most closely associated with low bone perfusion.
Hypothesis 2—Low post-debridement bone perfusion is associated with higher rates of 
infection.
3.0
Background*
Infection following high-energy trauma or blast injury is one of the most 
challenging complications facing musculoskeletal trauma patients, with an estimated 
annual cost of $35 billion in the US (https://stacks.cdc.gov/view/cdc/11550). Infection can 
be catastrophic, potentially causing prolonged morbidity, reduced function, permanent 
disability and amputation. Failed treatment results in chronic bone infection requiring 
repeat surgical procedures in approximately 30% of patients. 1-4
Blood supply plays a critical role in the bone health for trauma patients by 
delivering necessary oxygen, nutrients, antibiotics, and immune cells necessary to treat 
infection successfully, 5-9. The management of trauma-related infection is therefore based 
on aggressive, thorough debridement of all poorly perfused bone and connective tissue, 
followed by fracture stabilization to prevent further injury. More extensive debridement is 
believed to improve treatment success; however, additional tissue loss requires 
increasingly complex reconstructive procedures to fill bone and connective tissue voids. 
At present there are no objective, intraoperative, and real-time techniques to assess 
bone perfusion and thoroughness of debridement. This unmet clinical need results in 
substantial practice variation and places patients at unnecessarily high risk for chronic 
infection and potentially avoidable surgical procedures.
Our multidisciplinary research team has demonstrated that quantitative indocyanine 
green (ICG)-based Dynamic Contrast Enhanced Fluorescence Imaging (DCE-FI) can 
assess bone perfusion in a measurable, reproducible and predictable way using a porcine 
model.10 Furthermore, we have initiated an Institutional Review Board (IRB)-approved 
protocol to perform similar imaging in human trauma patients with good success. To this 
end, the primary objective of this project is to gather critical pilot data that will accelerate 
translation of ICG-based DCE-FI to clinical practice for human trauma patients. During 
this translational work we will systematically develop and evaluate an ICG-based DCE-FI 
methodology and analytic tools for intraoperative assessment of bone perfusion. Our long- 
term goal is to apply these sophisticated methods to guide surgical debridement that will 
enhance trauma patients’ short- and long-term recovery.

PROTOCOL TITLE: ICG Fluorescence Imaging in Lower Extremity Amputation Patients
4.0
Study Endpoints*
Primary outcome measure is ICG-based DCE-FI changes that occur as a result of 
osteotomy and soft tissue stripping associated with the amputation.
5.0
Study Intervention/Investigational Agent
5.1 Description:
ICG fluorescence imaging has been used to assess tissue perfusion in vivo in real-time in 
arterial and lymphatic perfusion imaging25, 26, osseous flap perfusion imaging27. ICG has 
been approved for clinical use by the FDA since 1959. The dynamic fluorescence 
imaging systems used in this study will be the Pentero surgical microscope (Zeiss, 
Germany),  SPY Elite (Novodaq/Stryker), or SPY PHI (Novodaq/Stryker). Both are 
publicly available imaging systems that can be used to assess tissue perfusion in real time 
in the operating room. The systems have a multi-directional imaging arm which contains a 
near-infrared light source that illuminates the fluorescent agent within the tissues, an HD 
video camera that captures the intensity of fluorescent marker in real-time, and software 
that allows the user to capture relative and absolute perfusion values within the surgical 
field. This product has been FDA approved for use to monitor blood flow, plastic surgery, 
microsurgery, reconstructive surgery, gastrointestinal imaging and coronary bypass 
surgery.
White
&NIR lights
Filter
ICG
ICG
(a)
This study is neither a drug nor a device trial. Patients 
will be administered FDA approved ICG through 
intravenous injection and imaged by a FDA approved 
surgical microscope (Pentero, SPY Elite, or SPY PHI) 
which is 0.5 meter away from the subject. Both ICG 
fluorescence and the two imaging systems have been 
used for routine clinical practice for many years. Figure 
(a) shows the Schematic sketch of the imaging systems. 
ICG fluorescence imaging utilizes intravenously injected 
ICG which is a fluorecent dy that is FDA-approved for 
clinical use, illuminated with near infrared light.  The 
ICG dye is indirectly activated and the dynamic 
fluorescence due to bone perfusion can be captured by a 
video rate imaging system.
Wh te 
lig
Lens

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This study is similar to STUDY00031575 in terms of methodology with a nearly 
identical intervention. That study is approved and active, but in a different patient 
population, both of which are highly clinically relevant.
5.2 Subject Compliance Monitoring
Subjects participate in a “one-time” event (i.e., surgery) as part of this study and are 
monitored through the in-patient surgical service during the time of their participation.
5.3 Receipt of Drug Supplies
Indocyanine green (ICG)/drape kits used in this study are purchased from Stryker. Each 
kit includes 25mg ICG, 10cc saline and a sterile drape. Kits will be delivered to DHMC 
Investigational Drug Services and to each relying site when needed who will count and 
verify that the shipment contains all of the items noted in the shipment inventory. Any 
damaged kits will be documented in study files and destroyed on site. This is standard of 
care for ICG/drape kits used clinically in non-research indications.
5.4 Storage
ICG will be stored in a locked temperature controlled room, as is standard of care for 
ICG/drape kits used clinically in non-research indications.
5.5 Dispensing of Study Drug
Once agent assignment is performed, research staff will transport ICG in its original 
package to the operating room. The surgeon or anesthesiologist will prepare the solution 
for injection per instructions from the Stryker. 0.1mg/kg ICG will then be administered 
per study protocol.
5.6 Drug Accountability
A member of the study team will document the amount of ICG administered and 
any amount remaining in the institution’s electronic medical record.
5.7 Return or Destruction of Study Drug
ICG is FDA approved for routine clinical practice. No return or destruction of study 
drug is needed for this study.
6.0
Procedures Involved*
This is a prospective non-randomized clinical trial which will assess bone perfusion using 

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ICG fluorescence imaging at several step in the surgical procedure to assess the effect of 
these steps on bone perfusion. This is neither a drug nor a device trial. Patients will be 
administered FDA approved ICG through intravenous injection and imaged using an FDA 
approved surgical microscope (Pentero,SPY Elite, or SPY PHI).
6.1 Patient selection
Consecutive patients 18 years of age or older who present to DHMC and are 
scheduled to undergo a lower extremity amputation (either below knee, through 
knee or above knee) will be considered for study enrollment.
6.2 Inclusion Criteria
The following criteria must be met by subjects to be eligible for study enrollment
a.
Patients 18 years of age or older
b. Scheduled for a lower extremity amputation (either below knee, 
through knee or above knee)
c.
Provision of informed consent
6.3 Exclusion Criteria
The following criteria would exclude a subject from study enrollment
a.
Iodine allergy
b. Pregnant or  breastfeeding women
6.4 Subject Recruitment and Retention
Participation in this research requires informed consent according to Institutional Review 
Board (IRB) guidelines and a signed IRB-approved Consent Form as the means of 
documenting this understanding. Potential recruits are instructed that their participation is 
completely voluntary and that their medical care will not be altered in any way should they 
elect not to participate at any time prior to surgery. Subjects are recruited from patients 
presenting or referred to Orthopedic Department at Dartmouth-Hitchcock Medical Center 
for treatment and meeting protocol inclusion criteria. Potential subjects will be approached 
by a member of their care team to gauge interest about participation in research prior to 
being approached by a member of the study team. Subjects will be invited to participate 
in this study by a member of the Orthopaedic Department, which will occur either at the 
time of consultation with the surgeon about the candidate’s standard-of-care procedures 
or at another time agreed to by the potential participant, the candidate’s surgeon and/or 
Dr. Gitajn and/or Dr. Henderson or their designee. No advertisements or other promotional 
material will be used. No finder fees or recruitment incentives will be offered. Women of 
child bearing potential are eligible for enrollment into this study because ICG 
administration is not considered to present any additional risk for these women. The 
study will exclude women who are pregnant or breast-feeding as indicated in the 
exclusion criteria. Women of child-bearing potential, if asked to participate, will be 

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given a pregnancy test at no cost to confirm pregnancy status before administration of 
ICG.
Eligible patients regardless of ethnicity or health status, will be identified and recruited 
subject to inclusion and exclusion criteria above. Patients who meet eligibility criteria will 
be asked to participate in the trial. If they agree written informed consent will be obtained 
from the patient or their healthcare proxy. To obtain informed consent, study personnel 
(surgeon or research coordinator) will adhere to the following procedures: (1) present study 
information in a manner that is understandable to the patient; (2) discuss the study with the 
patient and answer any questions; (3) allow the patient an opportunity to discuss 
participation with their family; (4) confirm that the patient understands the risks and 
benefits of participating in the study and that their participation is voluntary; (5) complete 
the consent process and obtain signatures from the patient and research team. The process 
of obtaining and documenting informed consent forms will be completed in accordance 
with local Good Clinical Practice (GCP) recommendations.
If the research team member obtaining consent is at all unsure about the patient's ability 
to consent, s/he will consult with the study PI.
A legally authorized representative (LAR) with reasonable knowledge of the potential 
participant will be approached to consent on the patient’s behalf if the patient cannot 
adequately answer at least 2 questions and it is determined that the patient’s level of 
cognition is not likely to change before surgery.
The choice of LAR will follow standard procedures. The following with be approached in 
this order of priority:
• Legal guardian
• Proxy (health care agent) named in an advance directive or durable power 
of attorney for health care
• Family member or other surrogate identified by the state law on health care 
decisions. Guidance will be provided to assist the LAR in making the 
consent decision. They will advised to base the decision on the 
participant’s expressed wishes, or, if these are not known, what they 
believe the participant would have desired under the circumstances of the 
injury, their beliefs and values. If the LAR does not know what the 
participant would have wanted, the LAR will advised to base the decision 
with the participant’s best interest in mind. They will be asked to carefully 
consider how much leeway the participant would likely give the LAR in 
making the choice about participation in the study.

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Recognizing that consent is an ongoing process, the study team will encourage the 
participants to ask additional questions that may arise during the course of their 
participation in the study.
6.5 Early Withdrawal of Subjects
Participants will be removed from the protocol if:
1) Study imaging is not completed for any reason. The subject withdraws consent.
2) The subject has an occurrence of a significant clinical event that precludes imaging.
3) The subject becomes pregnant.
If a participant is withdrawn from the protocol, the PI will mark the data of this subject as 
"withdrawn" and will add a detailed explanation about the cause of withdrawal in the 
database.
6.6 Study drug and device
6.6a Description
This study is neither a drug nor a device trial. Patients will be administered FDA 
approved ICG through intravenous injection and imaged by a FDA approved surgical 
microscope (Pentero,SPY Elite, or SPY PHI) which is 0.5 meter away from the subject. 
Both ICG fluorescence and the two imaging systems have been used for routine clinical 
practice for many years. Figure (a) shows the Schematic sketch of the imaging systems. 
ICG fluorescence imaging utilizes intravenously injected ICG, which is a fluorescent dye 
that is FDA-approved for clinical use, illuminated with near-infrared light.  The ICG dye 
is indirectly activated and the dynamic fluorescence due to bone perfusion can be 
captured by a video rate imaging system.
6.6.b Preparation and Administration of Study Drug
ICG used in this study is purchased from Novodaq/Stryker.
6.6.c ICG Preparation and Administration Procedures:
1) ICG is prepared by the Surgical and/or Anesthesia teams in the operating room 
on the day of surgery.
2) From the vial containing ICG, the required dose is withdrawn into a 
syringe, labeled, and administered through a peripheral intravenous line followed 
by a saline flush.
6.7 Study Procedures

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On the surgery day, the patient will be prepared and transported to surgery as per routine 
at Dartmouth-Hitchcock. In the OR, patient positioning, preparation of the surgical field, 
and draping will follow standard practice.
Prior to ICG administration, patients will undergo standard preoperative monitoring by the 
Same Day nursing staff that includes continuous pulse oximetry, blood pressure 
monitoring, and heart rate, and respiratory rate. These results will be available prior to 
surgery and prior to administration of ICG. ICG will be administered by an anesthetist in 
the operating room. Patients are under the care and observation of the anesthesia and 
surgical teams during this time. Postoperatively, patients are transferred to the Post- 
Anesthesia Care Unit (PACU) where they are monitored continuously by nursing staff 
supervised by anesthesiologists. During this entire time vital signs (including temperature, 
pulse, respiratory rate and blood pressure) are collected as standard of care. All clinical 
data will be reviewed for adverse events for a period of 30 minutes post-injection.
Day of surgery Procedures
Pre- 
Infusion
0-30
minutes 
post- 
infusion
Pulse
X
X
Blood Pressure
X
X
Temperature
X
X
Respiratory Rate
X
X
Adverse Events
X
A Pulse dye densometer (Pulsion Medical Systems), similar to a pulse oximetry probe, will 
be placed on the patient’s finger to acquire an arterial blood input function during ICG 
injection. After exposure of the tibia or femur 0.1 mg/kg ICG will be injected 
intravenously. Video rate ICG fluorescence images will be acquired 20 seconds before 
and 4 minutes after ICG injection. Osteotomy or bone cut will be performed and then 0.1 
mg/kg ICG will be injected intravenously and, again, video rate ICG fluorescence images 
will be acquired 20 seconds before and 4 minutes after ICG injection. Soft tissue stripping 
from tibia or femur will then be performed, per standard surgical procedure. Then a third 
dose of 0.1mg/kg ICG will be injected and video rate ICG fluorescence images will be 
acquired 20 seconds before and 4 minutes after ICG injection. There will be a minimum of 
5 minutes between ICG infusions to allow ICG to wash out of tissues.
Surgical plan will not be affected as a result of the intraoperative fluorescence imaging. 
Surgical care team will be blinded to the imaging results in the operating room. The 

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imaging screen will be either be covered or hidden from view of the surgical team. The 
operator of the imagine device may visualize the imaging screen to ensure integrity of 
imaging data collection but this person will not communicate the results to the surgical 
team.
There is no follow-up associated with this study.
7.0
Data and Specimen Banking*
Not applicable
8.0
Sharing of Results with Subjects*
This study is not intended to provide any results to the subjects because this is the 
investigational study and the subjects will be under standard practice for their 
condition and no diagnostic or therapeutic decisions will occur based on the study.
9.0
Study Timelines*
This study is a one-time imaging during the subject’s surgery. The duration of 
each subject enrolled in the study is from the date the subject signs the consent to 
the day that the surgery is carried out.
The duration to enroll all study subjects is Jan.2, 2020 – Dec.31, 2021.
The estimated date for the investigators to complete this study is Jan. 31, 2022
10.0
Inclusion and Exclusion Criteria*
Inclusion Criteria
a. Patients 18 years of age or older
b. Scheduled for a lower extremity amputation (either below knee, through knee 
or above knee)
c. Provision of informed consent
Exclusion Criteria
a. Iodine allergy
b. Pregnant women
c. Patients under the age of 18. 
d. Prisoners
e. Pregnant of nursing mothers

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Vulnerable Populations*
After the consent has been reviewed with a potential subject, the subject will be asked 
to tell the research team member two things about the research they recall. If they are 
not able to do so the subject will be considered incompetent to consent to the research 
trial. Patients found to be in capable of providing informed consent will be deemed by 
the research staff to have impaired decision making and therefore will only be 
consented if a LAR agrees to their participation and signs the consent form.
Patients with impaired decision-making capacity will be included in this study. If the 
patient is unable to provide informed consent (e.g. due to their injury) at the time they 
are identified, informed consent may be obtained from their legally authorized 
representative (LAR). Allowing informed consent from a patient’s LAR will reduce the 
risk of recruitment bias against the most severely infected patients.
For patient with impaired decision-making capacity, a LAR with knowledge of the 
potential participant will be approached to consent on the patient’s behalf. If the patient 
cannot adequately answer at least two questions and it is determined that the patient’s 
level of cognition is not likely to change before surgery, their LAR will be approached. 
The choice of LAR will follow standard procedures. The following will be approached 
in this order of priority:

legal guardian

proxy (health care agent) named in an advanced directive or durable power of 
attorney for health-care
The LAR will be advised to base the decision on the participant’s expressed wishes, or, if 
these are not known, what they believe the participant would have desired under the 
circumstances of the injury, their beliefs and values. Recognizing that consent is an 
ongoing process, the study team will encourage the participants to ask additional 
questions that may arise during the course of their participation in the study.
Subjects participation in this study completely voluntary. If at any time a patient finds 
participation to be unduly stressful the patient or their legal authorized representative 
may withdraw the patient from the study with no repercussions.
Assent will be obtained from all patients that are able to sign their name. If patients are 
injured so severely that they are not able to sign their name verbal assent will be obtained. 
Assent will not be obtained from patients whom are unconscious due to the nature of their 
injury. 
The study team will comply with consent procedures for cognitively impaired 
adults outlined in SOP HRP-013.

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11.0
Local Number of Subjects
The primary outcome measure is total bone blood flow (TBBF). Informed by data 
collected from the porcine study, a total sample size of 30 trans-tibial amputation 
patients is required to determine the difference in TBBF baseline bone perfusion 
kinetics and change in the bone perfusion kinetics associated with osteotomy and 
subsequent soft  tissue stripping. Considering that  the controlled environment 
associate with the porcine model may not be replicated to the same degree in the 
amputation sample, a more conservative difference of 0.84 standard deviations 
between baseline and osteotomy and soft tissue stripping was selected (i.e., 50% of 
the effect size identified in the porcine model). Assuming a one tailed test, a Type 
I error rate of 0.05, and a Type II error rate of 0.80, a sample of 20 was 
determined with an additional 10 amputation patients recruited in the event that 
there is any protocol- or data-related issues. Data analysis will be performed 
after half of the patients are enrolled to re-calculate sample size needed to 
determine a difference.
We anticipate that 20 subjects will be accrued locally. We anticipate that 20 
patients will be enrolled at Brigham and Women’s Hospital. A total of 40 patients 
will be enrolled.
12.0
Recruitment Methods 
Patients, 18 years of age or older who present to DHMC who are scheduled to 
undergo a lower extremity amputation (either below knee, through knee or above knee) 
will be considered for study enrollment. Participation in this research requires informed 
consent according to Institutional Review Board (IRB) guidelines and a signed IRB-
approved Consent Form as the means of documenting this understanding. This study 
requests a partial waiver of HIPAA for recruitment purposes. The partial waiver would allow for 
the surgeon to share with the research coordinator the patient name, sex, MRN number, age, 
underlying conditions as well as type and location of injury.  
A Potential subjects consenting to undergo a surgical procedure, will be informed by their 
surgeon or another member of their Orthopaedic care team that there is a research study 
for which they might be interested in participating.  Following this discussion, either at 
the same visit or a subsequent visit the patient will be approached by a member of the 
research study team who will present the study and inquire if the patient would like to 
participate.  Potential recruits are instructed that their participation is completely 
voluntary and that their medical care will not be altered in any way should they elect 
not to participate at any time prior to surgery. Following consent, inclusion and 

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exclusion criteria will be reviewed for all patients.  Patient who meet the appropriate 
eligibility criteria will be enrolled in the study.
No advertisements or other promotional material will be used. No finder fees or 
recruitment incentives will be offered. Women of child bearing potential are eligible for 
enrollment into this study because ICG administration is not considered to present any 
additional risk for these women. The study will exclude women who are pregnant or 
breast-feeding as indicated in the exclusion criteria. Women of child-bearing potential, if 
asked to participate, will be given a pregnancy test at no cost to confirm pregnancy 
status before administration of ICG.
13.0
Withdrawal of Subjects*
Participants will be removed from the protocol if:
1)
Study imaging is not completed for any reason.
2)
The subject withdraws consent.
3)
The subject has an occurrence of a significant clinical event that precludes imaging.
4)
The subject becomes pregnant.
If a participant is withdrawn from the protocol, the PI will mark the data of this subject 
as "withdrawn" and will add a detailed explanation about the cause of withdrawal in 
the database.
14.0
Risks to Subjects*
Risk of ICG injection: ICG (Indocyanine Green) will be administered intravenously 
three times: once after exposure, once after osteotomy and once after soft tissue 
stripping. ICG is FDA approved for human use in angiography for ophthalmology 
and cardiology applications and is given routinely to patients in these clinical 
settings. The risks are considered minimal and consist of nausea, vomiting, hives, 
increased heart rate in subjects with particular sensitivity to the dye. ICG does contain 
sodium iodide and patients with a history of allergy to iodides will be excluded. 
Anaphylactic or urticardial reactions are rare but have been reported in patients both 
with and without a history of allergy to iodides. Anaphylactic deaths have been 
reported following IGG administration during cardiac catheterization. Reported rates 
of mild, moderate and severe adverse reactions to ICG are 0.15%, 0.2% and 0.05%. 
Every effort will be made to minimize this risk as much as possible. It is the 
standard of care at this institution to obtain information related to allergies, 
sensitivities and past medical histories upon patient arrival. Patients will also be 
monitored throughout their hospitalization, as is the standard of care, by everyone 
involved in their medical care for evidence of new or previously unknown reactions or 
sensitivities. Additionally, at all times ICG fluorescence imaging assessments are taking 

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place, research staff and both surgical and anesthesia staff will monitor patients closely 
for any adverse reaction to ICG. Research staff and surgical and anesthesia staff will 
also monitor patients for at least 30 minutes after all ICG fluorescence assessment 
procedures have been completed. In the event of an unexpected allergic reaction, 
Dartmouth-Hitchcock Medical Center and all affiliated groups within this institution 
have procedures in place for managing patients with unknown or unexpected allergic 
reactions. If such a reaction were to occur, all standard of care treatments will be 
provided including but not limited to treatment with appropriate agents such as 
epinephrine, antihistamines and corticosteroids. Risk of infection from IV injection is 
also extremely rare but can occur. Pregnant women are excluded from the study. A 
pregnancy test is administered to women of child-bearing age on request at no cost.
Prolonged operating room time: Because the ICG fluorescence assessments will 
take place in the operating room, participation in this study may increase the amount 
of time spent under surgical anesthesia in the operating room. For the average case, an 
additional 10 minutes at most will be incurred. However, the research staff will make 
every effort to minimize this risk by performing the assessment test while other 
required operative procedures are being performed.
Risk of confidentiality breach: Subjects enrolled in research are always exposed 
to additional risks of a breach in confidentiality, for example, of some elements of 
their personal health information that is made available to study investigators as part 
of their participation. The risk is 100%, but the occurrence rate of an actual breach is < 
1% (to date, we are not aware of any subject participating in our studies who has 
experienced a detrimental breach of confidentiality or confidential information).
15.0
Potential Benefits to Subjects*
Patients enrolling in this study will not benefit directly because no diagnostic or 
therapeutic decisions will occur based on the study, and thus, administration of the study is 
not intended to alter the surgical procedure. However, future patients may benefit from 
the knowledge gained from the study since this study may provide an objective, 
intraoperative, and real- time methodology to assess bone perfusion and thoroughness of 
amputation.
16.0
Data Management* and Confidentiality
Confidentiality
Information about study subjects will be kept confidential and managed according to 
the requirements of the Health Insurance Portability and Accountability Act of 1996 
(HIPAA). Risk of breach of confidentiality of the medical records and status of 

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participants will be minimized. Databases which are used to store subject-sensitive 
information are password- protected and encrypted during file/data transfers from 
viewing terminals. Access will be limited to research team members who have 
undergone CPHS training at Dartmouth. Whenever possible and practical standard-of-
care clinical data used in the research will be de-identified when under analysis.
Case Report Forms
Study case report forms (CRFs) will be the primary data collection instruments for 
the study. All data requested on CRFs will be recorded. Any missing data will be 
explained. If a space on the CRF is left blank because the procedure was not performed or 
the question was not asked, a written notation will be made. If an item is not applicable to 
an individual case, written notation will be made. Changes to the CRFs will be initialed 
and dated.
Record Retention
Following closure of the study, the investigator will maintain all site study records in a safe 
and secure location. The records are maintained to allow easy and timely retrieval when 
needed (e.g., audit or inspection) and, whenever feasible, to allow any subsequent 
review of data in conjunction with assessment of the facility, supporting systems, and 
staff. Upon completion of study analysis, research information is stored in Dartmouth 
College Records Management off-site storage located at 6218 Etna Road, Hanover, NH 
03755. Documents are shredded on site after 50 years of storage.
17.0
Provisions to Monitor the Data to Ensure the Safety of Subjects*
This is a study with minimal risk
 
18.0
Provisions to Protect the Privacy Interests of Subjects
The information collected for this study will be used only for the purposes of research. It 
includes optical images, clinical images (i.e., CT, MRI), and surgical outcomes. 
Information on your gender, age, diagnosis, etc., will be collected in order to characterize 
some basic demographics of the subject population enrolled in the study. The data collected 
for this will be maintained indefinitely.
The identities of the people in this study will be carefully protected. Specifically, the 
subjects’ identity is coded into a study number and most of the data and analysis occurs 
with information that is archived only by study number. Information which identifies of 
the participants in the study is maintained in binders as required by the FDA, and these 

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binders will be kept in the office of research team. The office is kept locked when not in 
use, and the study team is trained in the importance of maintaining your confidentiality. 
Research data, if sent electronically to members of the research team, is de-identified and 
only secure file transfer methods are used.
The health information of the subjects can be only used by the research team member who 
is involved in the research. The research team includes the study director plus others 
working on this study at Dartmouth-Hitchcock Medical Center and elsewhere
The information collected for this study may be used by researchers or officials of 
the following institutions.
•
Dartmouth College
•
Mary Hitchcock Memorial Hospital
•
Dartmouth-Hitchcock Clinic
•
Dartmouth-Hitchcock Medical Center
Identifiable data collected for this study will be used for research purposes which 
are determined to be reasonable and in line with expectations by a review committee. 
Once data collected for this research study is no longer identifiable, the data may be 
used or disclosed for other purposes.
19.0
Compensation for Research-Related Injury
This is an imaging study with minimal risk. We do not anticipate any injury or sickness 
happen due to the study. If in any case subjects are injured or become ill as a result of 
research procedures, the subjects will be provided with medical treatment, but the 
following organizations do not plan to pay for this treatment.
Mary Hitchcock Memorial Hospital
Dartmouth-Hitchcock Clinic
Dartmouth-Hitchcock Medical Center
Trustees of Dartmouth College
20.0
Economic Burden to Subjects
Subjects will not be charged for any procedures conducted solely for research purposes, 
nor will they be charged for the SPY Elite® System (nor Pentero or SPY PHI) 
assessment.
Insurance plans will not be billed for research procedures that are not the usual care for 
subject’s condition. Some of the medical care that the subjects will receive during this 

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study is the usual care a doctor would recommend for subjects’ condition. Subjects or 
subjects’ insurance plan are expected to pay for the costs of this usual medical care.
21.0
Consent Process
To obtain informed consent, study personnel will adhere to the following procedures: 
(1) present study information in a manner that is understandable to the patient. a Spanish 
IRB approved translated copy of the consent will be used when appropriate (2) discuss 
the study with the patient and answer any questions; (3) allow the patient an 
opportunity to discuss participation with their family; (4) confirm that the patient 
understands the risks and benefits of participating in the study and that their 
participation is voluntary; and (5) complete the consent process and obtain signatures 
from the patient and research team. The process of obtaining and documenting 
informed consent will be completed in accordance with Good Clinical Practice. If the 
research team member obtaining consent is at all unsure about the patient's ability to 
consent, s/he will consult with the study PI.
Recognizing that consent is an ongoing process, the study team will encourage 
the participants to ask additional questions that may arise during the course of their 
participation in the study.
This study will not involve the subjects who is under age 18.
The study team will comply with consent procedures outlined in SOP HRP-090
Non-English Speaking Subjects
This study will consent patients for whom English is their second language.  This 
will pertain to only individuals whose native language is Spanish. Prospective 
subjects who present for traumatic amputation surgery will be consented with the 
assistance of a Spanish speaking representative from DHMC Interpreter Services. 
Spanish speaking individuals will be given the Informed consent that has been 
translated to Spanish to review and discuss via interpreter.  Subjects willing to 
participate will sign and date the Spanish version of the consent and be given a 
copy for their records.  The interpreter will sign the Spanish consent as a witness 
to the process.  Prospective subjects who present for traumatic amputation surgery 
will be 
22.0
Process to Document Consent in Writing
The consent form documentation process will follow SOP (HRP-091). This is an imaging 
study with minimal risk. The signed consent form will be uploaded to DHMC EDH.
Both the English version and the Spanish version of the consent form will be submitted 
for approval. 

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23.0
Setting
All consenting and imaging will be carried out in the orthopedic department or surgical 
unit in DHMC. The imaging processing will be carried out in Thayer biomedical 
imaging Lab located in Williamson 7F.
24.0
Resources Available
The Department of Orthopaedic Surgery at DHMC, chaired by David Jevsevar, MD, 
includes the Section of Orthopedics, Section of Physical Medicine and
Rehabilitation, the Spine Center, and the Center for Shared Decision Making.
Clinical activity consisted of approximately 55,000 ambulatory visits and 5,500 operative 
cases in 2014 with $215 million total revenue. Each new patient and surgical patient is 
tracked for appropriate care and patient reported outcomes (PROs). The effort led to a 
$24 million award from the Centers for Medicare and Medicaid Services (CMS) to create 
the High Value Health Care (HVHC) Innovation Center, to implement shared-decision-
making and PROs in 19 major medical centers across the US. DOS is distinguished 
nationally for research through the Spine Patient Outcomes Research Trial (SPORT) – 
the largest orthopedic surgery clinical trial ever funded by the NIH with over $30M from 
NIAMS since inception in 2001.
Dr. Gitajn is the principle investigator for the study.  Dr. Gitajn has multiple NIH and 
DoD funded projects. She is actively engaged in research in orthopaedic trauma surgery 
and clinical translation of early-stage medical devices and imaging systems.  
The Dynamic Surgical-Guidance Laboratory at Dartmouth-Hitchcock is directed by 
Assistant Professor Jonathan Elliott and is connected to Dartmouth-Hitchcock Medical 
Center, Norris Cotton Cancer Center, Center for Comparative Medicine and Research, 
Rubin Research Building and the state-of-the-art Williamson Translational Research 
Building by a shared hallway. Dr. Elliott leads a team of researchers including two 
postdocs, two graduate students, and a dedicated research engineer and is responsible for 
an annual research budget of over $1 million.
All study members, including clinicians, faculty members, staff and students, will be 
approved for this protocol before engaging in any activities related to this project, and 
will therefore be required to complete CITI training and be cleared by the Department of 
Orthopaedics before physically attending to any of the study procedures. 
25.0
Multi-Site Research*
This study will be conducted at two clinical sites,

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•
Dartmouth-Hitchcock Medical Center
•
Brigham and Women’s Hospital
We anticipate that 20 subjects will be accrued locally. We anticipate that 20 patients will 
be enrolled at Brigham and Women’s Hospital. A total of 40 patients will be enrolled.
This distribution of patients is subject to change during monitoring of rate enrollment at 
each clinical site.
All data will be communicated in a HIPPA-compliant manner using a HIPPA-compliant 
database for clinical data (REDCap) and a HIPPA-compliant storage drive for imaging 
data. Clinical and imaging data will be uploaded to the appropriate HIPPA-compliant 
site by the local research team at each institution and all data will be analyzed at either 
Dartmouth-Hitchcock Medical Center or Dartmouth College/Thayer School of 
Engineering. De-identified patient information using a Study ID number will be utilized 
when possible.