SARS-CoV-2 Seroconversion in Humans: A Detailed Protocol for a Serological Assay, Antigen Production, and Test Setup

Curr Protoc Microbiol. 2020 Jun;57(1):e100. doi: 10.1002/cpmc.100.


In late 2019, cases of atypical pneumonia were detected in China. The etiological agent was quickly identified as a betacoronavirus (named SARS-CoV-2), which has since caused a pandemic. Several methods allowing for the specific detection of viral nucleic acids have been established, but these only allow detection of the virus during a short period of time, generally during acute infection. Serological assays are urgently needed to conduct serosurveys, to understand the antibody responses mounted in response to the virus, and to identify individuals who are potentially immune to re-infection. Here we describe a detailed protocol for expression of antigens derived from the spike protein of SARS-CoV-2 that can serve as a substrate for immunological assays, as well as a two-stage serological enzyme-linked immunosorbent assay (ELISA). These assays can be used for research studies and for testing in clinical laboratories. © 2020 The Authors. Basic Protocol 1: Mammalian cell transfection and protein purification Basic Protocol 2: A two-stage ELISA for high-throughput screening of human serum samples for antibodies binding to the spike protein of SARS-CoV-2.

Keywords: COVID19; ELISA; SARS-CoV-2; protein expression; serological assay.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Antibodies, Viral / blood*
  • Antigens, Viral / biosynthesis*
  • Antigens, Viral / isolation & purification*
  • Betacoronavirus / immunology
  • COVID-19
  • COVID-19 Testing
  • Clinical Laboratory Techniques / methods*
  • Coronavirus Infections / blood
  • Coronavirus Infections / diagnosis*
  • Coronavirus Infections / immunology
  • HEK293 Cells
  • Humans
  • Pandemics
  • Pneumonia, Viral / blood
  • Pneumonia, Viral / diagnosis*
  • Pneumonia, Viral / immunology
  • Recombinant Proteins / immunology
  • Recombinant Proteins / isolation & purification
  • SARS-CoV-2
  • Seroconversion
  • Spike Glycoprotein, Coronavirus / biosynthesis*
  • Spike Glycoprotein, Coronavirus / genetics
  • Spike Glycoprotein, Coronavirus / immunology
  • Spike Glycoprotein, Coronavirus / isolation & purification*


  • Antibodies, Viral
  • Antigens, Viral
  • Recombinant Proteins
  • Spike Glycoprotein, Coronavirus
  • spike protein, SARS-CoV-2