tetano
Editor, Senior Moderator
Diagnostics (Basel)
. 2021 Oct 23;11(11):1970.
doi: 10.3390/diagnostics11111970.
Development of an Enzyme-Linked Immunosorbent Assay (ELISA) for Accurate and Prompt Coronavirus Disease 2019 (COVID-19) Diagnosis Using the Rational Selection of Serological Biomarkers
Theano Lagousi[SUP] 1 2 [/SUP], John Routsias[SUP] 3 [/SUP], Vana Spoulou[SUP] 1 2 [/SUP]
Affiliations
Abstract
Prompt COVID-19 diagnosis is urgently required to support infection control measures. Currently available serological tests for measuring SARS-CoV-2 antibodies use different target antigens, although their sensitivity and specificity presents a challenge. We aimed to develop an "in-house" serological ELISA to measure antibodies against SARS-CoV-2 by combining different protein antigens. Sera (n = 44) from COVID-19-confirmed patients were evaluated against different SARS-CoV-2 protein antigens and all potential combinations using ELISA. Patients' sera were also evaluated against commercially available ELISA diagnostic kits. The mixture containing RBD 2.5 μg/mL, S2 1 μg/mL and N 1.5 μg/mL was found to be the most potent. Plates were incubated with patients' sera (1:100), and goat anti-human alkaline phosphatase-conjugated IgG, ΙgM and IgA antibody was added. The cut-off value for each assay was determined using the mean optical density plus two standard deviations of pre-pandemic controls. The "in-house" ELISA displayed 91% sensitivity and 97% specificity for IgG antibodies, whereas its sensitivity and specificity for IgM and IgA were 75% and 95% and 73% and 91%, respectively. The "in-house" ELISA developed here combined three SARS-CoV-2 antigens (RBD, S2 and N) as capture antigens and displayed comparable and even higher sensitivity and specificity than otherwise quite reliable commercially available ELISA diagnostic kits.
Keywords: SARS-CoV-2; antibodies; serological tests.
. 2021 Oct 23;11(11):1970.
doi: 10.3390/diagnostics11111970.
Development of an Enzyme-Linked Immunosorbent Assay (ELISA) for Accurate and Prompt Coronavirus Disease 2019 (COVID-19) Diagnosis Using the Rational Selection of Serological Biomarkers
Theano Lagousi[SUP] 1 2 [/SUP], John Routsias[SUP] 3 [/SUP], Vana Spoulou[SUP] 1 2 [/SUP]
Affiliations
- PMID: 34829317
- DOI: 10.3390/diagnostics11111970
Abstract
Prompt COVID-19 diagnosis is urgently required to support infection control measures. Currently available serological tests for measuring SARS-CoV-2 antibodies use different target antigens, although their sensitivity and specificity presents a challenge. We aimed to develop an "in-house" serological ELISA to measure antibodies against SARS-CoV-2 by combining different protein antigens. Sera (n = 44) from COVID-19-confirmed patients were evaluated against different SARS-CoV-2 protein antigens and all potential combinations using ELISA. Patients' sera were also evaluated against commercially available ELISA diagnostic kits. The mixture containing RBD 2.5 μg/mL, S2 1 μg/mL and N 1.5 μg/mL was found to be the most potent. Plates were incubated with patients' sera (1:100), and goat anti-human alkaline phosphatase-conjugated IgG, ΙgM and IgA antibody was added. The cut-off value for each assay was determined using the mean optical density plus two standard deviations of pre-pandemic controls. The "in-house" ELISA displayed 91% sensitivity and 97% specificity for IgG antibodies, whereas its sensitivity and specificity for IgM and IgA were 75% and 95% and 73% and 91%, respectively. The "in-house" ELISA developed here combined three SARS-CoV-2 antigens (RBD, S2 and N) as capture antigens and displayed comparable and even higher sensitivity and specificity than otherwise quite reliable commercially available ELISA diagnostic kits.
Keywords: SARS-CoV-2; antibodies; serological tests.