COMPARISON OF DIRECT AND COVALENT GOLD NANOPARTICLE-IMMUNOGLOBULIN Y CONJUGATION IN LATERAL FLOW ASSAY FOR SARS-COV-2 RECEPTOR BINDING DOMAIN DETECTION

Abstract

Coronavirus 2019 (COVID-19) outbreak caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread globally, it has been a major impact to global public health and world economy. Therefore, large scale of screening test is needed to prevent the larger spread of COVID19. Lateral flow assay (LFA) is suitable for large populations screening due to its advantages such as inexpensive, rapid, and easy to use. As one of the main component of LFA, Gold nanoparticlesantibody conjugate has been rapidly explored as it affects the sensitivity of LFA which has been a main concern for a long time. Immunoglobulin Y (IgY) is an antibody that has good specificity because it does not interact with rheumatoid factors and protein A or G. Besides, IgY is easy to produce and has a high economic value so it is suitable for LFA biosensors. Direct and covalent conjugation methods have been widely used for gold nanoparticle-antibody conjugation. This research aim to compare gold nanoparticle-antibody conjugate using direct and covalent conjugation for detection of SARS-CoV-2 receptor binding domain (RBD). In this study, we first optimized various factors such as pH, buffer, and IgY concentration using spectrophotometry and naked eye for the direct conjugation, as for the covalent conjugation we optimized EDC/NHS concentration and linker (3-mercaptopropionic acid) concentration. Both conjugates were evaluated via lateral flow assay against RBD of SARS-CoV-2 and using ImageJ application. Our result shows that both methods has successfully detect the RBD of SARS-CoV- 2. We found that covalent conjugation method using 3-mercaptopropionic acid is more sensitive than direct conjugation respectively with an area of 8,438 and 1,721 using ImageJ area plot. This method comparison can be use as a insight for development of sensitive conjugate for LFA.