tetano
Editor, Senior Moderator
J Virol Methods
. 2024 Jul 5:114995.
doi: 10.1016/j.jviromet.2024.114995. Online ahead of print. A safe, cost-effective and high-throughput SARS-CoV-2 antigen capture ELISA suitable for large-scale screening in low-resource settings
Ashutosh Singh[SUP] 1 [/SUP], Upasana Sahu[SUP] 1 [/SUP], Pratik M Kulkarni[SUP] 2 [/SUP], Rupali Yadav[SUP] 1 [/SUP], Sandeep Bhatia[SUP] 1 [/SUP], H V Murugkar[SUP] 1 [/SUP], Madhusudan Hosamani[SUP] 2 [/SUP], Suresh Basagoudanavar[SUP] 2 [/SUP], Gaurav Kumar Sharma[SUP] 3 [/SUP], Praveen Kumar Gupta[SUP] 3 [/SUP], Naveen Kumar[SUP] 4 [/SUP], Aniket Sanyal[SUP] 1 [/SUP], Naveen Kumar[SUP] 5 [/SUP]
Affiliations
Diagnostics employing multiple modalities have been essential for controlling and managing COVID-19, caused by SARS-CoV-2. However, scaling up Reverse Transcription-Quantitative Polymerase Chain Reaction (RT-qPCR), the gold standard for SARS-CoV-2 detection, remains challenging in low and middle-income countries. Cost-effective and high-throughput alternatives like enzyme-linked immunosorbent assay (ELISA) could address this issue. We developed an in-house SARS-CoV-2 nucleocapsid capture ELISA, validated on 271 nasopharyngeal swab samples from humans (n = 252), bovines (n = 10), and dogs (n = 9). This ELISA has a detection limit of 195pg/100µL of N protein and does not cross-react with related coronaviruses, ensuring high specificity to SARS-CoV-2. Diagnostic performance was evaluated using ROC curve analysis, showing a diagnostic sensitivity of 67.78% and specificity of 100%. Sensitivity improved to 74.32% when excluding positive clinical samples with RT-qPCR Ct values > 25. Furthermore, inter-rater reliability analysis demonstrated substantial agreement (κ values = 0.73-0.80) with the VIRALDTECT II Multiplex RT-qPCR kit and perfect agreement with the CoVeasy[SUP]TM[/SUP] COVID-19 rapid antigen self-test (κ values = 0.89-0.93). Our findings demonstrated that the in-house nucleocapsid capture ELISA is suitable for SARS-CoV-2 testing in humans and animals, meeting the necessary sensitivity and specificity thresholds for cost-effective, large-scale screening.
Keywords: COVID-19 diagnostics; antigen capture ELISA; community screening; lateral flow assay; nucleocapsid protein.
. 2024 Jul 5:114995.
doi: 10.1016/j.jviromet.2024.114995. Online ahead of print. A safe, cost-effective and high-throughput SARS-CoV-2 antigen capture ELISA suitable for large-scale screening in low-resource settings
Ashutosh Singh[SUP] 1 [/SUP], Upasana Sahu[SUP] 1 [/SUP], Pratik M Kulkarni[SUP] 2 [/SUP], Rupali Yadav[SUP] 1 [/SUP], Sandeep Bhatia[SUP] 1 [/SUP], H V Murugkar[SUP] 1 [/SUP], Madhusudan Hosamani[SUP] 2 [/SUP], Suresh Basagoudanavar[SUP] 2 [/SUP], Gaurav Kumar Sharma[SUP] 3 [/SUP], Praveen Kumar Gupta[SUP] 3 [/SUP], Naveen Kumar[SUP] 4 [/SUP], Aniket Sanyal[SUP] 1 [/SUP], Naveen Kumar[SUP] 5 [/SUP]
Affiliations
- PMID: 38972641
- DOI: 10.1016/j.jviromet.2024.114995
Diagnostics employing multiple modalities have been essential for controlling and managing COVID-19, caused by SARS-CoV-2. However, scaling up Reverse Transcription-Quantitative Polymerase Chain Reaction (RT-qPCR), the gold standard for SARS-CoV-2 detection, remains challenging in low and middle-income countries. Cost-effective and high-throughput alternatives like enzyme-linked immunosorbent assay (ELISA) could address this issue. We developed an in-house SARS-CoV-2 nucleocapsid capture ELISA, validated on 271 nasopharyngeal swab samples from humans (n = 252), bovines (n = 10), and dogs (n = 9). This ELISA has a detection limit of 195pg/100µL of N protein and does not cross-react with related coronaviruses, ensuring high specificity to SARS-CoV-2. Diagnostic performance was evaluated using ROC curve analysis, showing a diagnostic sensitivity of 67.78% and specificity of 100%. Sensitivity improved to 74.32% when excluding positive clinical samples with RT-qPCR Ct values > 25. Furthermore, inter-rater reliability analysis demonstrated substantial agreement (κ values = 0.73-0.80) with the VIRALDTECT II Multiplex RT-qPCR kit and perfect agreement with the CoVeasy[SUP]TM[/SUP] COVID-19 rapid antigen self-test (κ values = 0.89-0.93). Our findings demonstrated that the in-house nucleocapsid capture ELISA is suitable for SARS-CoV-2 testing in humans and animals, meeting the necessary sensitivity and specificity thresholds for cost-effective, large-scale screening.
Keywords: COVID-19 diagnostics; antigen capture ELISA; community screening; lateral flow assay; nucleocapsid protein.