Diagnostics 2023, 13, 462. https://doi.org/10.3390/diagnostics13030462 www.mdpi.com/journal/diagnostics Article Use of Envelope Domain III Protein for the Detection of IgG Type Antibodies Specific to Zika Virus by Indirect ELISA Oumar Ndiaye 1,2 , Cheikh Tidiane Diagne 1 , Ahmed Abd El Wahed 3 , Fatou Dia 1 , Moussa Dia 1 , Adama Faye 2 , Silvania Da Veiga Leal 4 , Menilita dos Santos 4 , Maria da Luz de Lima Mendonça 4 , Carolina Cardoso da Silva Leite 5 , Cheikh Saad Bouh Boye 6 , Juliet E. Bryant 7 , Philippe Desprès 8 , Ousmane Faye 1 , Amadou Alpha Sall 1,† and Oumar Faye 1, * ,† 1 Virology Department, Institut Pasteur de Dakar, Dakar BP 220, Senegal 2 Institut de Santé et Développement (ISED), Université Cheikh Anta DIOP de Dakar, Dakar BP 16390, Senegal 3 Institute of Animal Hygiene and Veterinary Public Health, University of Leipzig, D‐04103 Leipzig, Germany 4 National Institute of Public Health, Largo do Desastre da Assistência, Chã de Areia, Praia P.O. Box 719, Cape Verde 5 World Health Organization, Country Office—AFRO Prédio Comum das Nações Unidas Meio Achada S. António 1° Andar, Praia P.O. Box 266, Cape Verde 6 Centre de Reference IST/VIH, Centre Hospitalier Universitaire Aristide Le Dantec, Dakar BP 3001, Senegal 7 The Global Fund Against AIDS, Tuberculosis and Malaria, Chemin du Pommier 40, 1218 Le Brand‐Sacconex, Switzerland 8 Processus Infectieux en Milieu Insulaire Tropical (PIMIT), Université de La Réunion, INSERM UMR 1187, CNRS 9192, IRD 249, Plateforme CYROI, 97490 Sainte‐Clotilde, France * Correspondence: oumar.faye@pasteur.sn; Tel.: +22‐133‐8399‐200 † These authors contributed equally to this work. Abstract: Zika virus (ZIKV) diagnostics are crucial for proper antenatal and postnatal care and also for surveillance and serosurvey studies. Since the viremia during ZIKV infection is fleeting, sero‐ logical testing is highly valuable to inform diagnosis. However, current serology tests using whole virus antigens frequently suffer from cross reactivity issues, delays, and technical complexity, espe‐ cially in low and middle income countries (LMICs) and endemic countries. Here, we describe an indirect ELISA to detect specific IgG antibodies using the ZIKV envelope domain III (EDIII) protein expressed in Drosophila S2 cells as an immunogen. Using a total of 367 clinical samples, we showed that the EDIII‐ELISA was able to detect IgG antibodies against ZIKV with high sensitivity of 100.0% and specificity of 94.7% when compared to plaque reduction neutralization tests (PRNTs) as the gold standard and using 0.208 as the cut‐off OD value. These results show the usefulness of the recombinant envelope domain III as an alternative to standard whole virus proteins for ZIKV diag‐ nostics as it improves the sensitivity and specificity of IgG ELISA assay when used as an immuno‐ gen. This method should, therefore, be extended to serological diagnostic techniques for other mem‐ bers of the flavivirus genus and for use in IgM diagnostic testing. Keywords: Zika; diagnostics; ELISA; recombinant; antigen; envelope; IgG 1. Introduction Zika virus (ZIKV) is an emerging arthropod borne virus transmitted by a number of mosquito species [1] belonging to the genus flavivirus of the family Flaviviridae. The first ZIKV outbreak was confirmed in Yapp Island in Micronesia in 2007, and throughout the last decade, it has been responsible of a number of outbreaks throughout the world [2]. The virus came to public attention in 2015–2016 when a dramatic increase in cases of Guil‐ lain Barré in adults and microcephaly cases in neonates in Brazil was linked to a large‐ Citation: Ndiaye, O.; Diagne, C.T.; Abd El Wahed, A.; Dia, F.; Dia, M.; Faye, A.; Leal, S.D.V.; dos Santos, M.; de Lima Mendonça, M.d.L.; da Silva Leite, C.C.; et al. Use of Envelope Domain III Protein for the Detection of IgG Type Antibodies Specific to Zika Virus by Indirect ELISA. Diagnostics 2023, 13, 462. https://doi.org/10.3390/ diagnostics13030462 Academic Editor: Laurent Bélec Received: 20 November 2022 Revised: 19 December 2022 Accepted: 23 December 2022 Published: 26 January 2023 Copyright: © 2023 by the authors. Li‐ censee MDPI, Basel, Switzerland. This article is an open access article distrib‐ uted under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecom‐ mons.org/licenses/by/4.0/).