A Safe Pseudotyped Lentivirus-Based Assay to Titer SARS-CoV-2 Neutralizing Antibodies
Measuring the neutralizing potential of SARS-CoV-2 antigens-exposed sera informs on effective humoral immunity. This is relevant to 1-monitor levels of protection within an asymptomatic population, 2-evaluate the efficacy of existing and novel vaccines against emerging variants, 3-test prospective therapeutic monoclonal neutralizing antibodies (NAbs) and, overall, to contribute to understand SARS-CoV-2 immunity. However, the goldstandard method to titer NAbs is a functional assay of virus-mediated infection, which requires biosafety level 3 (BSL-3) facilities. As these facilities are insufficient in Latin American countries, including Mexico, scant information has been obtained about NAb in these countries during the COVID-19 pandemic. An alternative solution to acquire NAb information locally is to use non-replicative viral particles that display the SARS-CoV-2 Spike (S) protein on their surface, and deliver a reporter gene into target cells upon transduction. Here we present the development of a NAb-measuring assay based on Nanolucmediated luminescence measurements from SARS-CoV-2 S-pseudotyped lentiviral particleinfected cells. The successive steps of development are presented, including lentiviral particles production, target cell selection, and TCID50 determination. We applied the optimized assay in a BSL-2 facility to measure NAbs in 15 pre-pandemic, 18 COVID-19 convalescent and 32 BNT162b2 vaccinated serum samples, which evidenced the assay with 100% sensitivity, 86.6% specificity and 96% accuracy. The assay highlighted heterogeneity in neutralization curves which are relevant in discussing neutralization potency dynamics. This is the first report of a safe functional assay to measure SARS-CoV-2 in Mexico and a cornerstone methodology necessary to measure NAb in the context of limited resources settings.Funding Information:We acknowledge the financial support received by CONACYT (scholarships 1007842 and 657487), Secretaría de Relaciones Exteriores, Tecnológico de Monterrey and StrainBiotech S.A de C.V, that made this work possible.Conflict of Interests: None to declare. Ethical Approval: Protocols for the use of human samples for this work were approved by the IRB of the Instituto Mexicano del Seguro Social (IMSS) with reference number R-2020-785-068 prior to starting this work.