(Invited) Enhancing Sensitivity of Lateral Flow Assays By Immobilization of Affinity Molecules in Hydrogel Polymer Embedded Test Lines

Abstract

Rapid tests of COVID-19 have played a critical role in controlling the virus spread during the pandemic. Lateral flow assays (LFA), a paper-based technique, are widely adopted as a platform for detecting SARS-CoV-2 antigens, anti-SARS-CoV-2 antibodies, and viral RNA in combination with other technologies such as CRISPR. Due to its rapidness, cheapness, and visual readout, LFA is especially suitable for the at-home test. However, LFA has manifested a low sensitivity for antigen-based COVID-19 test. It has been reported that the sensitivity was as low as 12–18% when Ct values of RT-PCR were between 30 and 37, which limits LFA as a diagnostic tool.We intend to develop a sensitive LFA for at-home use, like a pregnancy test strip, with a configuration shown in Figure 1a. Our first efforts are to lower its limit of detection (LoD) by enhancing the efficiency of molecular capture on the nitrocellulose membrane. As shown in Figure 1b, a test line on the strip comprises a hydrogel polymer, which would provide a biologically friendly environment for molecular interactions. We have fabricated a polyacrylamide test line containing streptavidin on a nitrocellulose strip using photolithography. Thus, a test strip can be produced by printing biotinylated affinity molecules on the test line. We have used gold nanoparticles as a colorimetric readout of LFA. Our results show that the immobilization of an antibody in the hydrogel polymer would enhance the sensitivity of LFA’s detection. Compared to those by physical adsorption of antibodies directly to the nitrocellulose membrane, our LFA reduced the LoD by ~ 2 orders of magnitude. We are currently applying our LFA to detect neutralizing antibodies and variants of concern (VOC) of SARS-CoV-2. In this presentation, we will report the current progress of our project. Figure 1. Illustration of (a) general configuration of lateral flow strips (b) a lateral flow strip with hydrogel polymer embedded in the test line for antibody testing Figure 1