tetano
Editor, Senior Moderator
Anal Chem
. 2024 Nov 7.
doi: 10.1021/acs.analchem.4c03574. Online ahead of print. A Novel Cell- and Virus-Free SARS-CoV-2 Neutralizing Antibody ELISA Based on Site-Specific Labeling Technology
Hongliang Liu[SUP] 1 2 [/SUP], Tiantian Liu[SUP] 1 2 [/SUP], Aiping Wang[SUP] 1 2 [/SUP], Chao Liang[SUP] 1 2 [/SUP], Xifang Zhu[SUP] 1 2 [/SUP], Jingming Zhou[SUP] 1 2 [/SUP], Yumei Chen[SUP] 1 2 [/SUP], Yankai Liu[SUP] 1 2 [/SUP], Yanhua Qi[SUP] 1 2 [/SUP], Wenjing Chen[SUP] 1 [/SUP], Gaiping Zhang[SUP] 1 3 2 [/SUP]
Affiliations
The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to the global spread of coronavirus disease 2019 (COVID-19), creating an urgent need for updated methods to evaluate immune responses to vaccines and therapeutic strategies. In this study, we introduce a novel cell-free, virus-free SARS-CoV-2 neutralizing antibody ELISA (NAb-ELISA), which is based on competitive inhibition of the receptor binding domain (RBD) of spike protein binding to the angiotensin-converting enzyme 2 (ACE2) receptor. In this method, site-specific biotinylated hACE2-Fc-Avi recombinant protein is immobilized onto a 96-well plate for capture, and the RBD-Fc-vHRP recombinant proteins serve as detection probes. Evaluation of sera from wild type (WT) or Delta RBD-immunized mice using the NAb-ELISA and pseudovirus neutralization tests (pVNTs) demonstrated strong correlations between assays (R[SUP]2[/SUP] = 0.91 and 0.90 for the WT and Delta groups, respectively). Additionally, the NAb-ELISA successfully detected cross-neutralizing activity in sera, though with slightly lower correlation to pVNT (R[SUP]2[/SUP] = 0.70-0.83). By employing NAb-ELISA instead of an indirect ELISA for hybridoma screening, five monoclonal antibodies (mAbs) with neutralizing activities against WT, Delta, and BA.2 pseudoviruses were obtained. This assay offers a straightforward, rapid, and safe approach to characterizing vaccine-induced antibody responses and mAb neutralization activity. Notably, the NAb-ELISA platform can be quickly adapted to assess neutralizing antibody responses against emerging mutant strains, addressing the rapid mutation of the virus.
. 2024 Nov 7.
doi: 10.1021/acs.analchem.4c03574. Online ahead of print. A Novel Cell- and Virus-Free SARS-CoV-2 Neutralizing Antibody ELISA Based on Site-Specific Labeling Technology
Hongliang Liu[SUP] 1 2 [/SUP], Tiantian Liu[SUP] 1 2 [/SUP], Aiping Wang[SUP] 1 2 [/SUP], Chao Liang[SUP] 1 2 [/SUP], Xifang Zhu[SUP] 1 2 [/SUP], Jingming Zhou[SUP] 1 2 [/SUP], Yumei Chen[SUP] 1 2 [/SUP], Yankai Liu[SUP] 1 2 [/SUP], Yanhua Qi[SUP] 1 2 [/SUP], Wenjing Chen[SUP] 1 [/SUP], Gaiping Zhang[SUP] 1 3 2 [/SUP]
Affiliations
- PMID: 39506608
- DOI: 10.1021/acs.analchem.4c03574
The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to the global spread of coronavirus disease 2019 (COVID-19), creating an urgent need for updated methods to evaluate immune responses to vaccines and therapeutic strategies. In this study, we introduce a novel cell-free, virus-free SARS-CoV-2 neutralizing antibody ELISA (NAb-ELISA), which is based on competitive inhibition of the receptor binding domain (RBD) of spike protein binding to the angiotensin-converting enzyme 2 (ACE2) receptor. In this method, site-specific biotinylated hACE2-Fc-Avi recombinant protein is immobilized onto a 96-well plate for capture, and the RBD-Fc-vHRP recombinant proteins serve as detection probes. Evaluation of sera from wild type (WT) or Delta RBD-immunized mice using the NAb-ELISA and pseudovirus neutralization tests (pVNTs) demonstrated strong correlations between assays (R[SUP]2[/SUP] = 0.91 and 0.90 for the WT and Delta groups, respectively). Additionally, the NAb-ELISA successfully detected cross-neutralizing activity in sera, though with slightly lower correlation to pVNT (R[SUP]2[/SUP] = 0.70-0.83). By employing NAb-ELISA instead of an indirect ELISA for hybridoma screening, five monoclonal antibodies (mAbs) with neutralizing activities against WT, Delta, and BA.2 pseudoviruses were obtained. This assay offers a straightforward, rapid, and safe approach to characterizing vaccine-induced antibody responses and mAb neutralization activity. Notably, the NAb-ELISA platform can be quickly adapted to assess neutralizing antibody responses against emerging mutant strains, addressing the rapid mutation of the virus.