The SARS-CoV-2 virus is the causing agent of the coronavirus disease 2019 (COVID-19) pandemic responsible for millions of deaths worldwide. The development of the humoral response to the virus has been the subject of intensive research. A flow cytometry-based assay using native full-length SARS-CoV-2 Spike protein expressed in 293 T cells was recently proposed as a complementary seropositivity assay. The aim of our study was to further develop the flow cytometry assay and to standardize its parameters for reliable inter-laboratory use. We have optimized the protocol, established the Receiving Operating Characteristic (ROC) curve and tested reproducibility using pre-COVID and convalescent, SARS-CoV-2 individual plasma samples. The flow-based assay was simplified and standardized by cultivating the 293 T cells in suspension and expressing results in Mean Equivalent Soluble Fluorochrome (MESF) using an internal antibody positive control. The ROC curve was determined with an area under the curve (AUC) of 0.996 and the assay specificity and sensitivity were established at 100% and 97.7% respectively. Reproducibility was good as determined on multiple cytometers, on different days, and with data acquisition as far as 72 h post-staining. The standardized assay could be used as a high throughput confirmatory assay in flow cytometry laboratories involved in serological testing.
Supplementary information: The online version contains supplementary material available at 10.1007/s10616-021-00511-1.
Keywords: Antibodies; Flow cytometry; SARS-CoV-2; Seropositivity testing; Spike; Standardization.
© The Author(s), under exclusive licence to Springer Nature B.V. 2021.