Development Of A Lateral Flow Assay For Sars-Cov-2 Nucleocapsid Protein Detection

Abstract

SARS-CoV-2 is a dangerous pandemic, threatening human life from 2019 until now. Dealing with the pandemic, lots of molecular biology detection methods had been introduced to detect the virus such as qRT-PCR, ELISA, LAMP, etc. However, the limitations of these methods are the long-time results and cross-infection. Therefore, lateral flow assay (LFA) had been developed as a highly potential point of care (POC) application, which is rapid, decentralized, and easy to interpret. Based on a sandwich immunoassay format, this study developed a lateral flow assay (LFA) targeting the nucleocapsid protein to detect the SARSCoV-2 virus at a laboratory scale. Detection Ab conjugates were prepared using AuNPs of 40 nm and 80 nm in size. Moreover, the application of horseradish peroxidase (HRP) and TMB substrate to enhance signal intensities was done to improve the detection limit of based AuNPs LFA, which could only reach 10ng/ml. By HRP enhancement, the limit of detection (LoD) of the based-AuNPs LFA was increased 20-fold from 10ng/ml to 500pg/mL of SARS-CoV-2 N protein in saliva. In addition, with the extraction of commercial buffer, the LoD of the homemade LFA had been increased 10-fold to 50pg/ml. Compared to other conventional antigen detection methods, the LFA combining HRP enhancement technique seems new, convenient, and straightforward to use and takes a brief time to complete with uncomplicated protocols. This study's results indicated that N-protein is a promising target to detect the SARS-CoV-2 virus by LFA applying the HRP enhancement