Evaluating the Neutralizing Immune Response in SARS-CoV-2 Infection By Serological Assays
A recent publication from E. Lau et al. in Nature Communications demonstrated that a SARS-CoV-2 infection elicits robust neutralizing antibody titers in most individuals that last beyond six months as confirmed using Chondrex, Inc.’s ChonBlock™ blocking/sample dilution buffer for ELISA.
To combat the COVID-19 pandemic, understanding the adaptive immune response, especially the neutralizing antibody response, is essential. However, given the short history of this disease, there are few reports on the subject. Evidence suggests that increased levels of neutralizing antibodies correlate to protection against SARS-CoV-2, hence collecting data that can elucidate the viral immune response is needed. Scientists collected sera from patients, confirmed SARS-CoV-2 infection with RT-PCR and quantified the responses of neutralizing antibodies by performing plaque reduction neutralization tests (PRNT). In addition, the duration of positive antibody responses to the SARS-CoV-2 spike protein receptor binding domain (RBD) were analyzed in ELISA and compared with the PRNT results.
Antibody levels against RBD were determined in 231 serum samples collected from 150 patients. 96 well plates were coated overnight with purified recombinant RBD protein in PBS buffer. The next day, plates were blocked with ChonBlock™ buffer and incubated for 2 hours at room temperature. Each sample was diluted at 1:100 with ChonBlock™ buffer, added in duplicate to the wells, and incubated for 2 hours at 37°C. After washing with PBS containing 0.1% Tween 20, horseradish peroxidase (HRP)-conjugated goat anti-human IgG was added to all wells and incubated for 1 hour at 37°C. The ELISA plates were then washed five times and TMB was added into each well. After a 15 minute incubation period, the reaction was stopped by adding 2 M H2SO4 and the absorbance was analyzed at a 450 nm wavelength. Extrapolation of the Weibull curve fitted to the aggregate data of severe, mild, and asymptomatic individuals suggests that it takes 448, 337, and 105 days respectively, for ELISA OD450 values to drop to the negative cut-off value. Longitudinal data from 134 serum samples from 54 patients estimated that the RBD ELISA values will be negative after approximately 495 and 416 days in severe and mild patients respectively.
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1. Lau, E.H.Y., Tsang, O.T.Y., Hui, D.S.C. et al. Neutralizing antibody titres in SARS-CoV-2 infections. Nat Commun 12, 63 (2021).
ChonBlock™ buffers are proprietary buffers which substantially reduce background noise reactions in ELISAs. ChonBlock™ Blocking and Sample Dilution Buffer is especially useful for assaying antibodies in human and animal sera at low dilutions due to its ability to block the non-specific binding of immunoglobulins in samples. Chondrex, Inc. also offers ChonBlock™ Detection Antibody Dilution Buffer for reducing background noise reactions caused by secondary antibodies. Please refer to the following links to Chondrex, Inc’s website for more information.