Methodology for evaluation of the level of IgG antibodies against different SARS-CoV-2 proteins using multiplex immunofluorescence analysis

Abstract

Introduction. The definition of patients’ serological status for SARS-CoV-2 is required for a clinical COVID-19 history evaluation and evaluation of epidemiological situation in general and also for development of population revaccination strategy. Though there are many different test systems for SARS-CoV-2 antibodies identifying, their sensitivity and specificity vary greatly. Test systems differ depending on used platform and antigen kit which make difficulties in the interpretation and comparison of results.

Aim – define diagnostic characteristics of MILLIPLEX® SARS-CoV-2 Antigen Panel 1 IgG test system for multiplex immunofluorescence assay of SARS-CoV-2 antibodies level on FlexMap 3D device (Luminex, USA) in blood serum samples of persons who have recovered from COVID-19 and those who were vaccinated against COVID-19.

Material and methods. The study includes 3 groups of participants: persons with history of COVID-19; persons without history of COVID-19 and unvaccinated; vaccinated ones. Identifying of the level of IgG antibodies against SARS-CoV-2 and virus neutralizing antibodies level was made for all samples using ELISA and multiplex immunofluorescence assay.

Results. In the course of this study a threshold value of IgG antibodies to SARS-CoV-2 S1 subunit of S protein (spike) was determined using multiplex immunofluorescence assay and it was 3267 MFI, also the sensitivity and accuracy of this method were assessed, which are 0.78 and 0.92 respectively. A linear regression model was developed for converting MFI values to BAU/ml values. Seroprotective level of antibodies to S1 subunit of SARS-CoV-2 S protein was determined and it was 5192 MFI.

Conclusion. Identifying of IgG antibodies using multiplex immunofluorescence assay has a high diagnostic sensitivity and specificity. This method allows to evaluate seroprotectiveness by level of antibodies to S1 subunit of SARS-CoV-2 S protein. Besides that, results could be compared to international units BAU/ml using formula:

m – 492,770 + 31,320 × b b = – 15,733 + 0,032 × m,

where b is a BAU/ml value, m is an MFI value.

Keywords:multiplex immunofluorescence assay; SARS-CoV-2; COVID-19; humoral immunity; IgG

For citation: Frolova L.V., Zemsky P.U., Mitrofanov S.I., Mukhin V.E., Shpakova T.A., Kazakova P.G., Akhmerova Yu.N., Bulanova N.V., Golubnikova L.A., Grammatikati K.S., Zhdanova A.S., Mkrtchian A.A., Sergeev A.P., Snigir E.A., Feliz N.V., Makarov V.V., Yudin V.S., Keskinov A.A., Kraevoy S.A., Yudin S.M., Skvortsova V.I. Methodology for evaluation of the level of IgG antibodies against different SARS-CoV-2 proteins using multiplex immunofluorescence analysis. Immunologiya. 2023; 44 (1): 109–19. DOI: https://doi.org/10.33029/0206-4952-2023-44-1-109-119 (in Russian)

Funding. The study had no sponsor support.

Conflict of interests. Authors declare no conflict of interests.

Authors’ contribution. Authors contributed equally.

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