Measurement of Oxygen Saturation in Healthy Human Volunteers Before, During, and After Hyperemic Events Using Multi-Modal Techniques: Spatial Frequency Domain Imaging, Transcutaneous Oxygen Measurement, Pulse Oximeter, and Apple Watch
Phase: NA
Conditions: Oxygen Saturation
Sponsor: Indiana University

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Version Date: 1.18.2022 
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Protocol Title 
 
Measurement of Oxygen Saturation in healthy human volunteers before, during and after 
hyperemic events using Multi-Modal Techniques: Spatial Frequency Domain Imaging (SFDI), 
transcutaneous oxygen measurement (TCOM), Pulse Oximeter, and Apple Watch. 
 
 
Principal Investigator: 
Surya C. Gnyawali, Ph.D. 
Assistant Professor, 
Indiana Center for Regenerative Medicine and Engineering at Indiana University 
Department of Surgery, Division of Plastic Surgery 
School of Medicine, Indiana University 
 
 
 
 
 
 
 
 
 
 

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Table of Contents: 
Study Schema 
1.0 
Background & Rationale 
2.0 
Objective(s) 
2.1 
Primary Objective 
2.2 
Secondary Objective 
2.3 
Tertiary/Exploratory/Correlative Objectives 
3.0 
Outcome Measures 
3.1 
Primary Outcome Measures 
3.2 
Secondary Outcome Measures 
3.3 
Tertiary/ Exploratory/ Correlative Outcome Measures 
4.0 
Eligibility Criteria 
4.1 
Inclusion Criteria 
4.2 
Exclusion Criteria 
5.0 
Study Design 
6.0 
Enrollment/Randomization 
7.0 
Study Procedures 
8.0 
Data Safety Monitoring 
9.0 
Study Withdrawal/Discontinuation 
10.0 
Statistical Considerations 
11.0 
Data Management 
12.0 
Privacy/Confidentiality Issues 
13.0 
Record Retention 
14.0 
References 

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 Background & Rationale 
Pulse oximeters are commonly utilized medical devices used to measure blood oxygen saturation (SpO2)[2]. 
These devices (such as the INNOVO Fingertip Pulse Oximeter) are either stand-alone or integrated into 
physiologic monitoring systems, using 2 wavelengths of lights to determine SpO2. With recent advances in 
technology, Spatial Frequency Domain Imaging (SFDI) using range of light wavelengths from red to near 
infrared (NIR) such as in Clarifi(R) (Modulated Imaging, Inc., Irvine CA), smartphones such as Apple Watch 
(Apple Inc., Cupertino, CA), transcutaneous oximetry TCOM (Perimed Inc., Las Vegas, NV) now have 
pulse oximetry capabilities. Since it is possible that most patients could utilize this technology, we sought 
to assess the accuracy, reliability and usability of these oximeters and compare outcomes. In this study, a 
cohort of 20 healthy volunteers above the age of 18 including males and females of different skin colors 
will be assessed at the same site and data will be compared. We aim to provide a set of data that will support 
the clinical and scientific community and identify more than one reliable skin oxygen measurement 
modalities.  The Innovo pulse oximeter device will be used for all subjects. 
1.0 
Objective(s) 
1.1 
Primary Objective 
To investigate multiplatform for tissue oxygen measurement devices 
1.2 
Secondary Objective 
To compare other modalities with FDA approved Pulse Oximetry 
1.3 
Tertiary/Exploratory/Correlative Objectives 
To investigate skin color biasness in the optical Oximeters. Fitzpatrick’s skin type chart 
will be used to evaluate skin type.  
2.0 
Outcome Measures/Endpoints 
2.1 
Primary Outcome Measures 
Oxygen in tissue and blood measured non-invasively 
2.2 
Secondary Outcome Measures 
Comparison of the devices provides the standard for clinical use 
2.3 
Tertiary/Exploratory/Correlative Outcome Measures 
Melanin or scarred tissue oxygenation compared to normal skin 
 
3.0 
Eligibility Criteria 
3.1 
Inclusion Criteria 
• 
Healthy volunteers 
• 
Able to understand and complete the Informed Consent  
• 
Both males and females 
• 
Age between 18-65 years 
• 
All ethnic backgrounds 
3.2 
Exclusion Criteria 
• 
Patients or limited health conditions 
• 
Under the age of 18 years or above the age of 65 years 
• 
Smoking tobacco product 
• 
Prisoners 
• 
Cannot consent for themselves 

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• 
Pregnant individuals 
4.0 
Study Design 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
5.0 
Enrollment/Randomization 
Potential subjects will be identified from IU School of Medicine staff members.  Each subject will use all 
four devices. 
6.0 
Study Procedures 
First the subjects will go through a questionnaire session about their medical history by a 
designated study team member. The questionnaire data will be placed in RedCap using a subject 
ID that cannot be linked back to the subject. Then the subject will be consented. Subject will be 
enrolled. Then the non-invasive data collecting procedure will start on the following equipment: 
Spatial Frequency Domain Imaging (SFDI): During consenting period, the Modulim equipment will be 
calibrated and setup ready for scanning. Subject’s volar aspect of the thumb along with the palm surface 
Recruitment of initial subject 
Initial screening and consenting 
Conduct a baseline assessment using 4 devices: Spatial 
Frequency Domain Imaging, Transcutaneous oxygen monitoring, 
Apple Watch Oxygen Sensor, and Pulse Oximeter for Oxygen 
Monitoring, then occlude the arm with 200 mmHg using blood 
pressure cuff for 2 minutes per device [1] and measure the 
oxygenation during and post occlusion.  
Analyze and interpret the data 

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will be ready to target the optical camera head. Actual scanning takes less than 60 seconds. The images will 
be processed offline. During processing 3-5 different regions of interest (ROI) will be taken to measure the 
oxygen parameters such as tissue oxygen saturation (StO2), oxy-hemoglobin (HbO2), deoxy-hemoglobin 
(HbR), superficial hemoglobin (HbT1, sub-surface hemoglobin (HbT2).   The model used is a Clarifi 
Modulum 
Transcutaneous oxygen monitoring (TCOM):  Transcutaneous oxygen monitoring (TCOM or TcpO2) is 
a noninvasive, clinically-approved method to obtain skin oxygen levels. The method is quantitative, and 
measures oxygen delivery to the skin from underlying tissue. Before positioning the electrode, an adhesive 
fixation ring will be placed on the dry skin on the volar aspect of the thumb and an electrolyte as a contact 
liquid will be filled half and the probe is aligned into it by rotating clock-wise to fasten it. Recording will 
be started and waited for the oxygen level to stabilize and a fixed value will be recorded.  The model used 
is a Perimed PeriFlux 5000.   The probe will be heated to about 45oC.  Although this device has other 
options, only the measurement of O2 will be performed using this device. 
Apple Watch Oxygen Sensor: Smart watch blood oxygen sensors also measure blood oxygen level in the 
tissue. Apple Watch Series 6 introduced this new feature for monitoring blood oxygen level using light emitting 
diodes (LEDs) at the back of apple watches. A low blood oxygen level can be indicative of a serious health issue 
that needs immediate attention. The apple watch is equipped with green, red, and infrared LEDs that shine 
light onto the blood vessels in the wrist, with photodiodes measuring the amount of light reflected back. Apple's 
algorithms use this information to calculate the color of the blood, which is an indication of how much oxygen 
is in the blood. Bright red blood is well oxygenated, while darker blood has less oxygen. This can measure blood 
oxygen levels between 70 and 100 percent. Most healthy people have blood oxygen levels that range from 95 to 
100 percent. The apple watch sensor will be positioned on the user’s preferred wrist. 
Pulse Oximeter for Oxygen Monitoring: The pulse oximeter enables transcutaneous monitoring of the 
oxygen saturation of hemoglobin in arterial blood (StO2). Pulse oximetry is so widely prevalent in medical 
care that it is often regarded as a fifth vital sign[3]. It is important to understand how the technology 
functions as well as its limitations. To recognize the settings in which pulse oximeter readings of oxygen 
saturation (SpO2), an understanding of two basic principles of pulse oximetry is required: (i) how 
oxyhemoglobin (HbO2) is distinguished from deoxyhemoglobin (HbR) and (ii) how the SpO2 is calculated 
only from the arterial compartment of blood. Pulse oximetry is based on the principle that HbO2 and HbR 
differentially absorb red and near-infrared (IR) light. It is fortuitous that HbO2 and HbR have significant 
differences in absorption at red and near-IR light because these two wavelengths penetrate tissues well 
whereas blue, green, yellow, and far-IR light are significantly absorbed by non-vascular tissues and water 
[3]. HbO2 absorbs greater amounts of IR light and lower amounts of red light than does HbR; this is 
consistent with experience – well-oxygenated blood with its higher concentrations of HbO2 appears bright 
red to the eye because it scatters more red light than does HbR. On the other hand, HHR absorb more red 
light and appears less red. Exploiting this difference in light absorption properties between HbO2 and HbR, 
pulse oximeters emit two wavelengths of light, red at 660 nm and near-IR at 940 nm from a pair of small 
light-emitting diodes located in one arm of the finger probe. The light that is transmitted through the finger 
is then detected by a photodiode on the opposite arm of the probe. In this study, volar aspect of thumb and 
index finder of the subject will be used to measure SpO2. The model is Innovo iP 900AP. 
Sequence of the events during the study visit: 
1) Consenting and enrolling the subjects 
2) Medical questionnaire 
3) Fitzpatrick’s skin type assessment 

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4) Non-invasive imaging and data collection of the thumb and index fingers with the above 
devices in the following order: 
a) Baseline imaging and data collection 
b) Occlusion of the blood flow using pressure cuff at 200mmHg for 2 minutes and then 
collect the data during occlusion. There is a minimal risk of pain on the arm inflation 
of the pressure cuff up to 200mmHg. Please see literature (1) where they have used 
240mmHg. 
c) Releasing the cuff and collecting the data during the hyperemic phase (post-deflation 
of the cuff). 
7.0 
Reportable Events 
Since all measurement modalities are non-invasive, there are no adverse events anticipated or unanticipated 
problems involving risk to participants.  The risk of the ring adhesive would be minimal as it could chafe 
the skin when removed.   
8.0 
Study Withdrawal/Discontinuation 
Participant can withdraw/discontinue himself or herself from the study. Since the study involves only one 
visit the process of withdrawal is not effective. However, participants can withdraw the study in any time 
during the visit.   
 
9.0 
Statistical Considerations 
Five sets of data will be collected from each of the devices and statistical mean and standard error 
of deviation from the mean will calculated.  
 
10.0 
Statistical Data Management 
Primary data will be collected via direct data measurement and imaging, whichever applies to the devices. 
TCOM, Pulse oximeter and apple watch date will be recorded with pen & paper and entered on the spread 
sheet and stored electronically in REDCap. The storage location will be backed up manually for every 
subject and at the end of the subject visit. The quality control methods will be used single entry with random 
checks of accuracy.  
11.0 
Privacy/Confidentiality Issues 
Regarding the method for ensuring participant privacy and for protecting privacy and confidentiality, each 
subject will be given a subject ID and only non-identifiable demographic data will be collected. Data will 
be stored on REDCap and access will be limited to designated personnel who processes the data. 

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12.0 
Follow-up and Record Retention 
The duration of the data collection will be 1-2 hours and the study will be within 6 months. The duration of 
record retention will be for 5 years and after that destruction or the possibility of indefinite archiving of 
information. 
13.0 
References 
 
1. 
Jones, M.D., J.L. Taylor, and B.K. Barry, Occlusion of blood flow attenuates exercise-
induced hypoalgesia in the occluded limb of healthy adults. J Appl Physiol (1985), 2017. 
122(5): p. 1284-1291. 
2. 
Nitzan, M., A. Romem, and R. Koppel, Pulse oximetry: fundamentals and technology 
update. Med Devices (Auckl), 2014. 7: p. 231-9. 
3. 
Chan, E.D., M.M. Chan, and M.M. Chan, Pulse oximetry: understanding its basic 
principles facilitates appreciation of its limitations. Respir Med, 2013. 107(6): p. 789-99.