Towards Quantitative and Standardized Serological and Neutralization Assays for COVID-19

Int J Mol Sci. 2021 Mar 8;22(5):2723. doi: 10.3390/ijms22052723.


Quantitative and robust serology assays are critical measurements underpinning global COVID-19 response to diagnostic, surveillance, and vaccine development. Here, we report a proof-of-concept approach for the development of quantitative, multiplexed flow cytometry-based serological and neutralization assays. The serology assays test the IgG and IgM against both the full-length spike antigens and the receptor binding domain (RBD) of the spike antigen. Benchmarking against an RBD-specific SARS-CoV IgG reference standard, the anti-SARS-CoV-2 RBD antibody titer was quantified in the range of 37.6 µg/mL to 31.0 ng/mL. The quantitative assays are highly specific with no correlative cross-reactivity with the spike proteins of MERS, SARS1, OC43 and HKU1 viruses. We further demonstrated good correlation between anti-RBD antibody titers and neutralizing antibody titers. The suite of serology and neutralization assays help to improve measurement confidence and are complementary and foundational for clinical and epidemiologic studies.

Keywords: IgG; IgM; RBD; SARS-CoV-2 virus; cross reactivity; monoclonal antibody reference standard; neutralization assay; quantitative serology assays; sensitivity and specificity; spike.

MeSH terms

  • Antibodies, Neutralizing / blood
  • Antibodies, Neutralizing / immunology
  • Antibodies, Viral / blood
  • Antibodies, Viral / immunology
  • COVID-19 / blood*
  • COVID-19 / immunology*
  • COVID-19 Serological Testing / methods*
  • COVID-19 Serological Testing / standards*
  • Cross Reactions
  • Flow Cytometry / methods
  • Fluorescence
  • Humans
  • Immunoglobulin G / blood
  • Immunoglobulin G / immunology
  • Immunoglobulin M / blood
  • Immunoglobulin M / immunology
  • Microspheres
  • Neutralization Tests / methods*
  • Neutralization Tests / standards*
  • Receptors, Virus / chemistry
  • Receptors, Virus / immunology
  • SARS-CoV-2 / immunology*
  • Spike Glycoprotein, Coronavirus / chemistry
  • Spike Glycoprotein, Coronavirus / immunology


  • Antibodies, Neutralizing
  • Antibodies, Viral
  • Immunoglobulin G
  • Immunoglobulin M
  • Receptors, Virus
  • Spike Glycoprotein, Coronavirus
  • spike protein, SARS-CoV-2