Rapid Turnaround, Home-based Saliva Testing for COVID-19
Phase: NA
Conditions: Covid19
Sponsor: Stanford University

NCT04568122 
 
August 23, 2021 
Rapid turnaround, home-based saliva testing for COVID-19  
 
-Study Design-  
 
 
Principal Investigator(s): Manu Prakash, Manisha Desai, and Euan Ashley 
Study Team: Adam Larson, Shailabh Kumar, Anesta Kothari, Haley Hedlin, Derek Boothroyd, 
Albee Ling, Ariadna Garcia, Bryan Bunning, Steven Hershman, Jeffrey Christle, Jessica 
Kennedy, Reed Sprague, Bec Osborne, Ruth O’Hara 
 
 
 
 
 
 
 
 

 
 
2 
     Contents 
 
 
 
 
Study Design Summary 
3
Study Population: 
4
Criteria for Hospitalized Patients 
4
Inclusion Criteria 
4
Exclusion Criteria: 
4
Criteria for High Risk and Positive Population 
4
Inclusion Criteria: 
4
Exclusion Criteria: 
4
Criteria for Low Risk Population 
5
Inclusion Criteria: 
5
Exclusion Criteria: 
5
Rationale 
6
Detailed Description of the Technology 
6
Instruments used with the Test: 
6
Study Design 
9
Figure 1. 
9
Aims 
10
Primary Feasibility Outcome: 
10
Secondary Feasibility Outcomes: 
10
Primary Validity Outcome: 
11
Statistical Considerations: 
11
General Considerations: 
11
Analysis Sets 
11
Analysis of the Feasibility Outcomes 
11
Analysis of the Validity Outcomes 
12
Handling of Missing Data 
12
Recruitment: 
12
Consent 
13
Test Protocol 
13
Data Acquired 
15

 
 
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Study Design Summary 
 
Study Type: 
Experimental 
Estimated Enrollment: 
3060 participants 
Observational Model: 
Cohort 
Time Perspective: 
Prospective 
Official Title: 
 
Estimated Study Start Date: 
September 2020 
Estimated Primary Completion Date: 
December 2021 
Estimated Study Completion Date: 
January 2022 
 
 

 
 
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Study Population: 
The study population consists of a diverse group of Stanford faculty, staff, and students, 
and the local Santa Clara County community and specific external sources with known 
positive saliva samples and DUA/MTA in place. More specifically, there are three sub-
populations to address our aims: 
1) (Positive population) Hospitalized patients (N=15)  
2) (High risk and positive population) People from Bay area community and external 
sites with known positive saliva samples and DUA/MTA in place; Those with a 
suspected COVID infection seeking testing within the Stanford Health Care 
system (N=1500)  
3) (Low risk population) People from Bay area community; Students on the Stanford 
campus and employees at Stanford Health Care (N=1500)  
 
Criteria for Hospitalized Patients 
Inclusion Criteria 
● >=18 years 
● Hospitalized at Stanford Health Care for COVID-related complications and tested 
positive for SARS-CoV-2 with a PCR-based test 
● Able to understand and consent to study and with a clinical trajectory likely to be 
consistent with multi-day participation 
Exclusion Criteria: 
● Participants with salivary gland dysfunction (including patients with Sjogren’s 
disease or those with xerostomia associated with lupus or rheumatoid arthritis) 
● Participants will not be eligible if they identify any reason they are unable to 
participate in the study 
Criteria for High Risk and Positive Population  
Inclusion Criteria: 
● >=18 years 
● Seeking testing for suspected COVID or a participant in a study of COVID-
positive outpatients or from external sites with known positive saliva samples and 
DUA/MTA in place 
● Own an internet-enabled phone or device capable of loading web pages, 
receiving text messages, and taking/uploading photos. 
● Willing to participate in the study for one month 

 
 
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Exclusion Criteria: 
● Participants who have color blindness 
● Participants unable to operate the SnapDx device 
● Participants with salivary gland dysfunction (including patients with Sjogren’s 
disease or those with xerostomia associated with lupus or rheumatoid arthritis) 
● Participants will not be eligible if they identify any reason they are unable to 
participate in the study 
Criteria for Low Risk Population 
Inclusion Criteria: 
● >=2 years 
● Have access to an internet-enabled phone or device capable of loading web 
pages, receiving text messages, and taking/uploading photos. 
● Willing to participate in the study for one month, or permission and willingness 
from legal guardian for participants under 18 years old.  
Exclusion Criteria: 
● Participants with prior confirmed SARS-CoV-2 infection 
● Participants who have color blindness 
● Participants unable to operate the SnapDx device 
● Participants with salivary gland dysfunction (including patients with Sjogren’s 
disease or those with xerostomia associated with lupus or rheumatoid arthritis) 
● Participants will not be eligible if they identify any reason they are unable to 
participate in the study 
Study Location: Stanford University Campus, Stanford Health Care, Stanford 
affiliated COVID-19 testing sites (including drive through testing sites). We will accept 
deidentified and inactivated saliva samples from external sources with DUA/ MTA in 
place and IRB approval.  
 
Funding: Stanford Medicine Catalyst 
 
DUA , MTA OTL: We have DUA in place with a site in Guatemala and will have 
DUA/ MTA in place for the sharing of saliva samples from any/all sources as they 
become available. 
 

 
 
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Mobile Medical Website/App: REDCap  
DRA: N/A or previously approved for Pattern Health 
 
IND Exempt: Yes, observational diagnostic tool 
 
Manufacturer: N/A 
 
Rationale 
Clinical testing for SARS-CoV-2 falls broadly into two categories: molecular testing to detect the 
genomic sequence of the virus and serology testing for the antibody response of the host. While 
CLIA based testing in both categories has proved valuable for individual care during the 
pandemic, the cost (up to $100 in some cases) and time (a minimum of 12 hours in most cases) 
required for these approaches mean they have had little public health impact on the pandemic. 
A home based surveillance test for the presence of SARS-CoV-2 in saliva, that is cheap (a few 
dollars) and fast (under 40 minutes) would be transformative in the global management of 
COVID-19. It would allow nothing less than the reopening of schools, businesses, and the safe 
return of many aspects of society to normal. Individuals could test themselves every morning 
before heading to work, for example, to demonstrate they were not currently infected. In this 
protocol, we aim to demonstrate the viability of such an approach in a target population of 
Stanford employees and students who would test themselves at our research facility and upload 
the results via a mobile website/app. The protocol includes enrichment for positive samples 
through a cohort recruited through the Stanford Emergency Department (that would allow 
calculations of sensitivity required by the US FDA) and CLIA validation of these samples, a 
collection of known Sars-Cov-2 samples from external sources (including from Guatemala) as 
well a random selection of negative home tests to allow estimation of specificity  The overall aim 
is to demonstrate the feasibility, usability, and real-world validity of home surveillance testing at 
scale to provide the foundation for scaling up to potentially millions of tests around the world. 
Detailed Description of the Technology 
The loop-mediated isothermal amplification (LAMP) assay has proven to be a reliable and 
simple protocol that can detect small amounts of viral RNA in patient samples. This LAMP 
protocol has the benefits of being simple, requiring no specialized equipment; rapid, requiring 
less than an hour from sample collection to readout; and cheap, costing around $1 per reaction 
using commercial reagents. Heating to separate the reaction inhibitors from inactivated samples 
was shown to be an effective way to ensure reliable LAMP amplification. In this protocol, we 
take advantage of an open hardware solution invented by one of the PIs (SnapDx) that can be 
assembled with readily available components for the cost of <$5 dollars a unit and could be 
used together with the LAMP assay for point of care detection of COVID-19 RNA from saliva. 
We further utilize a novel substrate - cellulose - for RNA extraction to further simplify the 
protocol for LAMP. Finally, we utilize a novel approach in construction of the assay to run the 

 
 
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home tests in a manner to eliminate any chances for reaction cross-contamination; common in 
laboratory molecular tests.  
 
Instruments used with the Test: 
The SnapDx is designed to be a low-cost at-home device for detecting SARS-CoV-2 in saliva 
samples. The process utilizes LAMP (isothermal amplification) reagents to detect SARS-CoV-2 
in saliva. With COVID-19 cases increasing globally, diagnostics need to be accessible to 
everyone, including resource-limited regions. 
SnapDx test hardware: $3-5 dollars per unit (estimate) 
Saliva tube and solutions: $1 dollar per unit (estimate) 
Reagents and Materials: 
Cost breakdown for reagents: 
● TCEP - 1G (Pierce 20490) >3000 reactions - $80 
● EDTA - 100mL (Sigma 03690) >10,000 reactions - $40 
● Primers - (IDT) 1.2ml of 10X >500 reactions - $30 
● NEB master mix (E1700L) >500 reactions - $650 
 
Primers: It is good practice to make 20µL 100µM aliquots once they are resuspended. In case 
of contamination a fresh stock can be used. For a 10X stock- 16 μl FIP, 16 μl BIP, 2 μl F3, 2 μl 
B3, 4 μl LF, 4 μl LB, and brought to 100 μl with water. 
Ordered from IDT, sequences from Rabe 2020. 
As1_F3 
CGGTGGACAAATTGTCAC 
As1_B3 
CTTCTCTGGATTTAACACACTT 
As1_LF 
TTACAAGCTTAAAGAATGTCTGAACACT 
As1_LB 
TTGAATTTAGGTGAAACATTTGTCACG 
As1e_FIP 

 
 
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TCAGCACACAAAGCCAAAAATTTATTTTTCTGTGCAAAGGAAATTAAGGAG 
As1e_BIP 
TATTGGTGGAGCTAAACTTAAAGCCTTTTCTGTACAATCCCTTTGAGTG 
 
Materials included in SnapDx Kit: 
 
Item 
Manufacturer 
Order Number 
Spit tube 
BIC  
 
SnapDx 
BIC  
 
Instructions for Use  
PrakashLab 
 
 
 
 
Materials included in the COVID-19 LAMP diagnostic assay (Gold Standard - clinically 
indicated): 
 
Same as above.  
Item 
Manufacturer 
Order Number 
LAMP master mix 
NEB 
M1804 
primers 
IDT 
Standard Desalting 
 

 
 
9 
 
Study Design 
 
 
Figure 1  
Study flowchart for recruitment, assay performance, and follow-up. Blue and yellow icons 
indicate processes and data collection respectively, in which key steps are colored with darker 
shades. CRC = Clinical Research Coordinator, PRO = patient reported outcome, MRN = 
medical record number. 
 
Usability/Feasibility 
All study procedures involving contact with individuals of any infectious disease risk status (low, 
moderate, high) will be performed by trained clinical research personnel with all the necessary 
personal protective equipment. All of these personnel have also been vaccinated against 
COVID-19. Nonetheless they will use standard precautions, including getting tested weekly and 
staying away from others including study personnel and potential participants, in any case of 
symptoms or illness.  
 
A cohort of 120 participants with diverse demographic backgrounds will be recruited and each of 
them will take one SnapDx test at our study site. Additional saliva samples will be collected to 
be used for laboratory testing for clinical evaluation purposes. Participants will complete a 
Usability Survey towards the end of taking the SnapDx on our study site to answer questions on 
satisfaction, ease, information, and use. These Usability questions include the following with 
multiple choice answers (strongly disagree, disagree, somewhat disagree, neither agree nor 
disagree, somewhat agree, agree, strongly agree): 

 
 
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- 
(Satisfaction) Overall, I am satisfied with how easy it was to use the test.  
- 
(Ease) It was easy to learn to use this test. 
- 
(Information) It was easy to find the information I needed. 
- 
(Use) If the SnapDx test were available to me today, I would use this on a regular basis. 
- 
We also ask If they would be willing to retake the test 
 
Answers to these questions will be summarized and reported to the FDA as the results of our 
Human Usability Study. 
 
Validity 
Saliva samples are to be collected in several patient populations, including from both 
inpatient and outpatient clinics, and COVID positive or negative patients determined by nasal 
swab PCR tests from several different locations. We will make a concerted effort to reach out to 
the Stanford and bay area communities and collect positive saliva samples from those who 
have tested positive through a standard PCR test. Five intended users will be given basic 
training to conduct SnapDx tests on these saliva samples as if they were taking the test 
themselves. A small portion of each saliva sample will also be tested for SARS-COV-2 using 
saliva direct PCR in the laboratory. SnapDx test results will be compared against saliva direct 
PCR results to determine the positive percent agreement (PPA) and negative percent 
agreement (NPA). 
 
Aims 
The aim of the study is to demonstrate the feasibility and validity of a saliva based home 
surveillance monitoring test for SARS-CoV-2 infection. These will be addressed in the 
following sub-aims. 
1) To assess whether there is sufficient concordance between the technician and 
patient in interpretation of findings (Primary Feasibility Aim)  
2) To assess sensitivity and specificity of the home saliva test (Primary Validity Aim)  
 
Primary Feasibility Outcome:  
An indicator for discrepancy in interpretation of findings by participant versus by 
technician via photo (interrater reliability) 
 
Secondary Feasibility Outcomes:  
There are 4 secondary outcomes involved in assessing feasibility. These include 
 

 
 
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1) An indicator for ambiguous findings as measured by percent confidence (if < 80% 
confident) in interpretation of results by participant and by technician (ease of 
interpretation)  
2) An indicator that the participant appropriately followed instructions for using the 
kit (≥ 80% confident) (ease of use) 
3) An indicator for whether participant called technical support and by reason (e.g., 
device leaked or there was a possible broken part) (ease of use) 
4) An indicator by participant that the sample was sufficient and reached the 
indicated line for sample integrity (sample integrity)  
 
Primary Validity Outcome:  
An indicator for whether the home saliva test is positive (to assess sensitivity and 
specificity) 
 
Statistical Considerations: 
 
General Considerations: 
We will use the Agresti-Coull method (Agresti and Coull 1998) to construct confidence 
intervals. Unless otherwise noted, all confidence intervals constructed will be 95% 
confidence intervals. 
 
Analysis Sets: 
There are two key analysis sets that are subsets of the study population. The first 
subset consists of outpatients. The second subset consists of the patients hospitalized 
for COVID-related complications who will be tested daily for 5 days. 
 
Analysis of the Feasibility Outcomes 
Analyses of all feasibility outcomes will be performed in the outpatient analysis set. 
 
The concordance of the test results will be compared to the results of PCR testing. We 
will calculate the proportion in agreement (# of tests where SnapDx and PCR testing 
agree/# total tests), the proportion correctly identified as positive (# of tests SnapDx-
positive/# PCR-positive), and the proportion correctly identified as negative (# of tests 
SnapDx-negative/# PCR-negative). A threshold of 80% overall agreement and 85% 
agreement in the positive tests will indicate the home test is feasible. 
 

 
 
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Percent confidence in interpretation of the results and that instructions were followed 
correctly will be summarized as 1) percent of respondents stating that they are at least 
80% confident in their interpretation and 2) median and interquartile range of reported 
confidence levels. Results will be reported separately for participants and technicians. 
 
The proportion of participants who call technical support will be presented and the 
frequency of the reason for the call will be presented. The proportion of tests that have a 
sufficient amount of saliva to reach the indicator line will be presented. 
 
The feasibility outcomes will be presented separately by demographic characteristics in 
a subgroup. For example, we will evaluate whether the concordance is comparable 
across different age groups, between men and women, etc. 
 
Analysis of the Validity Outcomes 
The sensitivity, specificity, positive predictive value (PPV), and negative predictive value 
(NPV) of the test will be estimated using the results collected in outpatients. The results 
of the PCR-based test will be the gold standard used to estimate the test’s diagnostic 
accuracy. Home test results will only be included in this analysis if the test is performed 
within 24 hours of the time the swab used in the PCR tests was collected. 
 
Handling of Missing Data 
Although we have designed our study to minimize the amount of missing data, we 
expect to have a small amount of missing data. For example, hospitalized patients who 
are very sick or are discharged prior to day 5 may not provide data for days 1-5; 
participants may not complete the surveys each time they provide results from their 
home tests; or some PCR test results may not be able to be linked to the home test 
results. Analyses in the outpatient population will assume that data is missing at random 
and will use multiple imputation methods to account for the missing data. Given the 
small number of hospitalized patients expected, we will perform a complete case 
analysis in the analysis set consisting of hospitalized patients. Sensitivity analyses will 
be performed to evaluate the robustness of our findings to assumptions about missing 
data.   
 
 
 
Recruitment: 
The study will recruit up to 3060 participants including 60 hospitalized patients from 
multiple locations. These will include SHC COVID testing sites, Stanford Health Care 

 
 
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300 and 500P locations (the Emergency Department and other in-patient wards 
particularly M7, the COVID ward), as well as the front entrance of Stanford hospital and 
several locations on Campus. We will also be recruiting folks from the bay area 
community for our study. Participants will fall into two categories: i) those receiving a 
CLIA certified nasal swab COVID-19 test at SHC at the moment of recruitment, and ii) 
those not.  
 
We will be recruiting a high-risk population because we will be recruiting from a pool 
that includes people who are seeking medical attention and/or participating in COVID-
related research studies. Feasibility and validity of the tests will be evaluated in the 
high-risk population. We will be additionally recruiting a low-risk population to assess 
feasibility in the general population. Hospitalized patients will provide information on 
concordance between the CLIA COVID test and the home saliva test in patients over a 
five day period when we expect to see a mix of positive and negative tests. 
 
To increase the number of SARS-CoV-2 positive saliva samples, we will accept 
deidentified and deactivated saliva samples from external sources, including 
Guatemala. All of these external sources will have DUA/MTA in place and will have 
institutional approval and have received written informed consent from all participants 
for the local trials, including the sharing of these samples with Stanford University for 
the current trial.  
 
Inclusion and Exclusion criteria:  
(as noted above) 
 
Consent 
Potential participants will be approached by dedicated clinical research coordinators 
and study volunteers. They will be asked if they have an interest in participating and if 
so, whether they have an internet data-enabled cell phone capable of uploading high 
resolution photographs. Interested participants will then be directed to the digital e-
consent implemented in REDCap on the mobile web browser. The CRC will be 
available to answer questions. The participants will also be provided an email and 
contact phone information for questions.  
 
For the pediatric population, informed consent will be given by the legal guardian after 
they have been given the opportunity to answer any and all of the questions they may 
have and assent will be provided for children able to understand and complete the assent 
form (generally at least 7 years old).  
 

 
 
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Consent and Assent form will include a request to share access to their medical records 
for review of the results of any CLIA certified COVID test and any medical condition that 
might affect the veracity of testing. It will also include a request to observe the 
participants in-person or via video conferencing. 
 
After consent, participants will receive a bag with materials for multiple home based 
tests as well as instructions on their phone via REDCap on how to use the test.  
 
Test Protocol 
The REDCap survey will include a video demonstration of the test procedure. In 
addition, written instructions for use (IFU) will be available.  
 
Participants will be asked to carry out as many tests as are included in the bag (most 
likely between one and two) on a daily basis until they are used up. 
 
Test description 
 
Test materials: saliva collection tube, small handheld centrifuge, test kit (plasticware) 
The test kit should be stored in a regular fridge (4C) at the location of the participant - 
prior to use.  
 
The test materials include a cartridge containing a tube with closed reagents into which 
the participant will provide the saliva sample. Participants will be asked to provide 1ml of 
saliva into the collection tube, after which they will close the cap of the cartridge. They 
will be asked to pour boiling water into a mug and place the tube into the boiling water 
for 3 minutes. At the end of 3 minutes, they will then place the tube back in the mug of 
water for 40 minutes. At the end of 40 minutes, they remove the tube and note the color 
(pink/ orange = negative, yellow = positive). They will take a photograph of the tube 
against a given white photocard (provided in the kit) and upload it to the website. After 
filling in the short (<1 minute) symptom questionnaire and the short usability (<20 
seconds) questionnaire they will dispose of the tube in the trash. 
 
At that point, if the test result is negative (no evidence of infection) then they will be 
asked to do nothing other than fill in a survey and upload the image of the tube via study 
app. If the test result is positive they will be instructed to behave as if they might be 
infectious in their home and outside of their home (maximizing the use of masks and 
distancing) and asked to take a CLIA nasal swab test at their nearest SHC testing site 
at their earliest opportunity.  
 

 
 
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Our study provides the option for the participants to take a PCR nasal swab test at Stanford 
Health Care when their SnapDx tests result or the saliva direct PCR test done by the lab shows 
a positive SARS-CoV-2 result. Our study will cover the cost of the PCR nasal swab test for the 
participants.  
 
 
Figure: Color change in tubes. Yellow color is a positive result  
 
 
 
 
 
 
Data Acquired 
Data acquired on all participants in this study will include 
1) A baseline demographics survey collecting age, sex, and race/ethnicity, 
education levels, socio-economic background and COVID-19 vaccination status 
2) A symptom survey filled in by the participant (one per test) 
 
Additional data acquired on the positive/at-risk and the low-risk participants includes: 
3) A survey to collect confidence in interpretation and in following the instructions 
(one per test) 
4) An associated image of the at-home LAMP assay test result (one per test) 
5) A follow-up survey for the usability comments (once) 
 
Data collected on participants who have a CLIA COVID-19 test performed within 24 
hours of the home saliva test will additionally include: 
1. A medical chart review for the results of any CLIA COVID-19 

 
 
16 
2. Technicians will additionally provide an evaluation of their confidence in 
interpreting the participant-provided images using a survey identical to the one 
shown to participants 
3. Also included will be the additional documentation for the participant to follow the 
local healthcare authorities and CDC guidelines for COVID-19 related process.  
 
 
Payments: 
Participants will not be paid for participation. The study team will cover the costs of any 
follow-up CLIA certified testing required.