{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,14]],"date-time":"2026-10-14T02:56:27Z","timestamp":1791946587722,"version":"4.3.4"},"reference-count":70,"publisher":"MDPI AG","issue":"1","license":[{"start":{"date-parts":[[2024,1,18]],"date-time":"2024-01-18T00:00:00Z","timestamp":1705536000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Bio-Manguinhos\/Oswaldo Cruz Foundation"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Diseases"],"abstract":"<jats:p>Massive vaccination positively impacted the SARS-CoV-2 pandemic, being a strategy to increase the titers of neutralizing antibodies (NAbs) in the population. Assessing NAb levels and understanding the kinetics of NAb responses is critical for evaluating immune protection. In this study, we optimized and validated a PRNT50 assay to assess 50% virus neutralization and evaluated its accuracy to measure NAbs to the original strain or variant of SARS-CoV-2. The optimal settings were selected, such as the cell (2 \u00d7 105 cells\/well) and CMC (1.5%) concentrations and the viral input (~60 PFU\/well) for PRNT-SARS-CoV-2 with cut-off point = 1.64 log5 based on the ROC curve (AUC = 0.999). The validated PRNT-SARS-CoV-2 assay presented high accuracy with an intraassay precision of 100% for testing samples with different NAb levels (low, medium, and high titers). The method displays high selectivity without cross-reactivity with dengue (DENV), measles (MV), zika (ZIKV), and yellow fever (YFV) viruses. In addition, the standardized PRNT-SARS-CoV-2 assay presented robustness when submitted to controlled variations. The validated PRNT assay was employed to test over 1000 specimens from subjects with positive or negative diagnoses for SARS-CoV-2 infection. Patients with severe COVID-19 exhibited higher levels of NAbs than those presenting mild symptoms for both the Wuhan strain and Omicron. In conclusion, this study provides a detailed description of an optimized and validated PRNT50 assay to monitor immune protection and to subsidize surveillance policies applied to epidemiologic studies of COVID-19.<\/jats:p>","DOI":"10.3390\/diseases12010029","type":"journal-article","created":{"date-parts":[[2024,1,18]],"date-time":"2024-01-18T03:50:56Z","timestamp":1705549856000},"page":"29","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":18,"title":["Plaque Reduction Neutralization Test (PRNT) Accuracy in Evaluating Humoral Immune Response to SARS-CoV-2"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0889-6991","authenticated-orcid":false,"given":"Ingrid Siciliano","family":"Horbach","sequence":"first","affiliation":[{"name":"Laborat\u00f3rio de An\u00e1lise Imunomolecular, Instituto de Tecnologia em Imunobiol\u00f3gicos Bio-Manguinhos, Funda\u00e7\u00e3o Oswaldo Cruz, Rio de Janeiro 21040-900, Brazil"},{"name":"Programa de P\u00f3s-Gradua\u00e7\u00e3o em Medicina Tropical, Instituto Oswaldo Cruz, Funda\u00e7\u00e3o Oswaldo Cruz, Rio de Janeiro 21040-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7401-5107","authenticated-orcid":false,"given":"Adriana","family":"de Souza Azevedo","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de An\u00e1lise Imunomolecular, Instituto de Tecnologia em Imunobiol\u00f3gicos Bio-Manguinhos, Funda\u00e7\u00e3o Oswaldo Cruz, Rio de Janeiro 21040-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Waleska Dias","family":"Schwarcz","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de An\u00e1lise Imunomolecular, Instituto de Tecnologia em Imunobiol\u00f3gicos Bio-Manguinhos, Funda\u00e7\u00e3o Oswaldo Cruz, Rio de Janeiro 21040-900, 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