Development of a Rapid Focus Reduction Neutralization Test Assay for Measuring SARS-CoV-2 Neutralizing Antibodies.

Vanderheiden, Abigail; Edara, Venkata Viswanadh; Floyd, Katharine; Kauffman, Robert C; Mantus, Grace; Anderson, Evan; Rouphael, Nadine; Edupuganti, Sri; Shi, Pei-Yong; Menachery, Vineet D; Wrammert, Jens; Suthar, Mehul S

SARS-CoV-2 is a recently emerged human coronavirus that has escalated to a pandemic. There are currently no approved vaccines for SARS-CoV-2, which causes severe respiratory illness or death. Defining the antibody response to SARS-CoV-2 will be essential for understanding disease progression, long-term immunity, and vaccine efficacy. Here we describe two methods for evaluating the neutralization capacity of SARS-CoV-2 antibodies. The basic protocol is a focus reduction neutralization test (FRNT), which involves immunostaining infected cells with a chromogen deposit readout. The alternate protocol is a modification of the FRNT that uses an infectious clone-derived SARS-CoV-2 virus expressing a fluorescent reporter. These protocols are adapted for use in a high-throughput setting, and are compatible with large-scale vaccine studies or clinical testing. © 2020 Wiley Periodicals LLC Basic Protocol: Focus reduction neutralization test Alternate Protocol: mNeonGreen-based focus reduction neutralization test (FRNT-mNG).