DiagnosDisk - a highly sensitive point-of-care test for detecting SARS-CoV-2 antigen in saliva

Project Summary/Abstract SARS-CoV-2, a global pandemic, has caused more than 6 million infections and 180,000 deaths in the United States alone during the past 7 months. Both the WHO and CDC emphasize the need for point-of-care (POC) tests to expand diagnosis of acute infections to control the pandemic (i.e. isolating infected individuals). The current gold standard test for diagnosing acute SARS-CoV-2 infection is reverse transcriptase polymerase chain reaction (RT-PCR) with nasopharyngeal samples. The assay is highly sensitive, but requires well-trained personnel for sample collection and locations with specialized equipment. Therefore, the turnaround time varies from one to few days, delaying isolation, contact tracing, and medical attention. These challenges can be addressed with lateral flow immunoassays (LFIA) to detect viral antigens in self-collected saliva samples. Compared to nasopharyngeal specimens, studies have demonstrated that assays with saliva samples are more consistent and the saliva itself may contain higher viral load. However, existing LFIA for detecting salivary antigens exhibit poor sensitivity (≤ 40%) caused by the use of small sample volumes and sample dilution. To achieve timely and accurate screening of suspected COVID-19 cases, this project aims to develop a rapid assay for detecting SARS-CoV-2 antigens in saliva with high sensitivity to meet WHO's target product profile. We hypothesize that using ≥500 µL saliva as the assay sample can significantly improve the sensitivity because a larger specimen volume provides more analytes for detection. The new test will employ temperature-responsive polymer-antibody conjugates in conjunction with DiagnosDisk, a novel flow-through assay device. The approach is innovative in in using enriched saliva specimens for more consistent assay results, enhancing antigen detection by scaling up the saliva specimen volume, and adopting the well- developed sandwich immunoassay for DiagnosDisk with temperature-responsive polymer reagents that simplify the design while maximizing the sensitivity. We will employ polymer-antibody conjugates that efficiently isolate analytes in saliva (Aim 1) and fabricate the DiagnosDisk device to accommodate larger volume saliva specimens to improve the detection limit (Aim 2). The sensitivity of the new rapid assay, which combines polymer-antibody conjugates and DiagnosDisk, will be evaluated using saliva specimens from COVID-19 positive and negative patients (Aim 3).