A pseudotyped lentivirus-based assay to titer SARS-CoV-2 neutralizing antibodies in Mexico

Cruz-Cardenas, J. A.; Gutierrez-Mayret, M.; Lopez-Arredondo, A.; Castaneda-Delgado, J. E.; Rojas-Martinez, A.; Garcia-Rivas, G.; Enciso-Moreno, J. A.; Palomares, L. A.; Brunck, M. E. G.

doi:10.1101/2022.01.27.478128

bioRxiv · 10.1101/2022.01.27.478128

A pseudotyped lentivirus-based assay to titer SARS-CoV-2 neutralizing antibodies in Mexico

Cruz-Cardenas, J. A.·Gutierrez-Mayret, M.·Lopez-Arredondo, A.·Castaneda-Delgado, J. E.·Rojas-Martinez, A.·Garcia-Rivas, G.·Enciso-Moreno, J. A.·Palomares, L. A.·Brunck, M. E. G.

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Abstract

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 gold-standard 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 Nanoluc-mediated luminescence measurements from SARS-CoV-2 S-pseudotyped lentiviral particle-infected 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. Overall, this is the first report of a BSL-2 safe functional assay to measure SARS-CoV-2 in Mexico and a cornerstone methodology necessary to measure NAb with a functional assay in the context of limited resources settings. ImportanceEvaluating effective humoral immunity against SARS-CoV-2 requires a functional assay with infectious virus. Handling the authentic SARS-CoV-2 virus requires specialized facilities that are not readily available in Latin America, including Mexico. Here we produce non-replicative viral particles pseudotyped with the SARS-CoV-2 S protein that are used as safe surrogate viral particles in an optimized BSL-2 ready neutralization assay. The establishment of this assay is critical to allow the evaluation of effective humoral immunity to SARS-CoV-2 post-infection and to monitor the efficacy of existing or novel vaccines against emerging variants in the Mexican population.

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Cruz-Cardenas, J. A., Gutierrez-Mayret, M., Lopez-Arredondo, A., Castaneda-Delgado, J. E., Rojas-Martinez, A., Garcia-Rivas, G., Enciso-Moreno, J. A., Palomares, L. A., Brunck, M. E. G.. 2022-02-02. A pseudotyped lentivirus-based assay to titer SARS-CoV-2 neutralizing antibodies in Mexico. https://doi.org/10.1101/2022.01.27.478128

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