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Multiplexed Flow Cytometric Approach for Detection of Anti-SARS-CoV-2 IgG, IgM and IgA Using Beads Covalently Coupled to the Nucleocapsid Protein

preprint
submitted on 16.03.2021, 01:09 and posted on 17.03.2021, 05:31 by Ingrid Fatima Zattoni, Luciano F. Huergo, Edileusa C. M. Gerhardt, Jeanine M. Nardin, Alexia Marques Fernandes dos Santos, Fabiane Gomes de Moraes Rego, Geraldo Picheth, Vivian Rotuno Moure, Glaucio Valdameri

Flow cytometry has emerged as a promising technique for detection of SARS-CoV-2 antibodies. In this study, we described a new methodology to detect simultaneously IgG, IgM and IgA of SARS-CoV-2 nucleocapsid protein in human serum by flow cytometry. The Nucleocapsid protein was covalently bound on functional beads surface applying sulfo-SMCC chemistry. BUV395 anti-IgG, BB515 anti-IgM, biotinylated anti-IgA1/IgA2 and BV421 streptavidin were used as fluorophore conjugated secondary antibodies. Serum and antibodies reaction conditions were optimized for each antibody isotype detection and a multiplexed detection assay was developed. This new cell-free multiplex approach based on flow cytometry was able to efficiently discriminate COVID-19 negative and positive samples. The simultaneous detection of IgG, IgM and IgA showed a sensibility of 88.5-96.2% and specificity of 100%. The combined detection of antibody isotypes offers greater spectrum for detection and monitoring of COVID-19 vaccines and seroconversion. This novel strategy opens a new avenue for flow cytometry-based diagnosis.

History

Email Address of Submitting Author

gvaldameri@ufpr.br

Institution

Federal University of Parana

Country

Brazil

ORCID For Submitting Author

0000-0003-1882-1512

Declaration of Conflict of Interest

No conflict of interest

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ChemRxiv

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