{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,25]],"date-time":"2026-10-25T03:30:27Z","timestamp":1792899027931,"version":"4.5.0"},"reference-count":29,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,16]],"date-time":"2026-06-16T00:00:00Z","timestamp":1781568000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100016302","name":"Coalition for Epidemic Preparedness Innovations (CEPI), Oslo, Norway","doi-asserted-by":"publisher","award":["PRJ-7355"],"award-info":[{"award-number":["PRJ-7355"]}],"id":[{"id":"10.13039\/100016302","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Background\/Objectives: Nipah virus (NiV) is a highly pathogenic zoonotic virus with fatality rates exceeding 70% and causes recurring outbreaks in South and Southeast Asia. Reliable serological assays are critical for outbreak surveillance, diagnosis, and evaluation of vaccine-induced immune responses. This study aimed to develop and qualify an indirect enzyme-linked immunosorbent assay (ELISA) based on recombinant NiV glycoprotein G for the detection of virus-specific IgG antibodies in human serum. Methods: An indirect ELISA was developed and optimized for antigen concentration, blocking conditions, and serum dilution. The assay performance was evaluated using convalescent human sera from Bangladesh, along with the World Health Organization (WHO) International Standard for anti-Nipah virus antibodies, maintained and distributed by the National Institute for Biological Standards and Control (NIBSC). Analytical validation was conducted in accordance with ICH Q2 (R2) guidelines, including assessments of sensitivity, specificity, Precision, Linearity, and detection limits. Results: The assay demonstrated 100% sensitivity and specificity relative to reference sera. Intra-assay coefficients of variation ranged from 0.36% to 5.73%, and inter-assay variation was 4.16%, indicating high precision. The ELISA showed excellent Linearity (R2 &gt; 0.995). The lower limit of detection was 0.51 IU\/mL, and the lower limit of quantification was 0.98 IU\/mL. Conclusions: The developed ELISA is a BSL-2-compatible, robust, and scalable platform suitable for serosurveillance and the assessment of vaccine-induced immunity in endemic regions. Calibration against an international standard supports its applicability for standardized antibody measurement. This assay provides a practical tool for NiV outbreak response and vaccine evaluation.<\/jats:p>","DOI":"10.3390\/vaccines14060534","type":"journal-article","created":{"date-parts":[[2026,6,17]],"date-time":"2026-06-17T03:27:21Z","timestamp":1781666841000},"page":"534","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Development and Qualification of a Nipah Virus Glycoprotein-Specific IgG ELISA for the Assessment of Human Antibody Responses"],"prefix":"10.3390","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0064-1812","authenticated-orcid":false,"given":"Mohammad Mamun","family":"Alam","sequence":"first","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-8184-790X","authenticated-orcid":false,"given":"Tahsin Tabassum","family":"Anonto","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sinthia","family":"Karim","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gathoni","family":"Kamuyu","sequence":"additional","affiliation":[{"name":"Coalition for Epidemic Preparedness Innovations (CEPI), 6th Floor, Henry Wood House, 4-5 Langham Place, London W1B 3DG, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-8394-4769","authenticated-orcid":false,"given":"Ali","family":"Azizi","sequence":"additional","affiliation":[{"name":"Coalition for Epidemic Preparedness Innovations (CEPI), 1899 Pennsylvania Avenue NW, Washington, DC 20006, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ayesha","family":"Siddika","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4939-4779","authenticated-orcid":false,"given":"Shadman Sakib","family":"Choudhury","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Md Wasik","family":"Rahman","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-4397-9653","authenticated-orcid":false,"given":"Anika","family":"Farzin","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-2337-6766","authenticated-orcid":false,"given":"Dewan Imtiaz","family":"Rahman","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8116-4511","authenticated-orcid":false,"given":"Rubhana","family":"Raqib","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6876-0191","authenticated-orcid":false,"given":"Mustafizur","family":"Rahman","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sharmin","family":"Sultana","sequence":"additional","affiliation":[{"name":"Institute of Epidemiology, Disease Control and Research (IEDCR), Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Trevor","family":"Shoemaker","sequence":"additional","affiliation":[{"name":"Viral Special Pathogens Branch, Division of High-Consequence Pathogens and Pathology, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, 1600 Clifton Rd. NE, Atlanta, GA 30333, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0409-7896","authenticated-orcid":false,"given":"Michael K.","family":"Lo","sequence":"additional","affiliation":[{"name":"Viral Special Pathogens Branch, Division of High-Consequence Pathogens and Pathology, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, 1600 Clifton Rd. NE, Atlanta, GA 30333, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5121-1962","authenticated-orcid":false,"given":"Sayera","family":"Banu","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5061-3783","authenticated-orcid":false,"given":"Tahmina","family":"Shirin","sequence":"additional","affiliation":[{"name":"Institute of Epidemiology, Disease Control and Research (IEDCR), Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christina F.","family":"Spiropoulou","sequence":"additional","affiliation":[{"name":"Viral Special Pathogens Branch, Division of High-Consequence Pathogens and Pathology, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, 1600 Clifton Rd. NE, Atlanta, GA 30333, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joel M.","family":"Montgomery","sequence":"additional","affiliation":[{"name":"Viral Special Pathogens Branch, Division of High-Consequence Pathogens and Pathology, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, 1600 Clifton Rd. NE, Atlanta, GA 30333, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2959-4522","authenticated-orcid":false,"given":"Syed Moinuddin","family":"Satter","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4103-4835","authenticated-orcid":false,"given":"Mohammed Ziaur","family":"Rahman","sequence":"additional","affiliation":[{"name":"One Health Laboratory, Infectious Diseases Division, International Centre for Diarrhoeal Disease Research, Mohakhali, Dhaka 1212, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,16]]},"reference":[{"key":"ref_1","first-page":"25","article-title":"Epidemiology of Henipavirus Disease in Humans","volume":"359","author":"Luby","year":"2012","journal-title":"Curr. Top. Microbiol. Immunol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"108263","DOI":"10.1016\/j.ijid.2025.108263","article-title":"Insights into the Acute Phase of Nipah Virus Infection: Clinical features, viral detection, and humoral immune response","volume":"163","author":"Satter","year":"2026","journal-title":"Int. J. Infect. Dis."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1432","DOI":"10.1126\/science.288.5470.1432","article-title":"Nipah Virus: A Recently Emergent Deadly Paramyxovirus","volume":"288","author":"Chua","year":"2000","journal-title":"Science"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"97","DOI":"10.1016\/j.coviro.2016.12.003","article-title":"Nipah Virus: Transmission of a Zoonotic Paramyxovirus","volume":"22","author":"Clayton","year":"2017","journal-title":"Curr. Opin. Virol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"2082","DOI":"10.3201\/eid1012.040701","article-title":"Nipah Virus Encephalitis Reemergence, Bangladesh","volume":"10","author":"Hsu","year":"2004","journal-title":"Emerg. Infect. Dis."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"e707","DOI":"10.1016\/S1473-3099(24)00262-7","article-title":"Measures to Prevent and Treat Nipah Virus Disease: Research Priorities for 2024\u201329","volume":"24","author":"Moore","year":"2024","journal-title":"Lancet Infect. Dis."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Fischer, K., Diederich, S., Smith, G., Reiche, S., Pinho dos Reis, V., Stroh, E., Groschup, M.H., Weingartl, H.M., and Balkema-Buschmann, A. (2018). Indirect ELISA Based on Hendra and Nipah Virus Proteins for the Detection of Henipavirus Specific Antibodies in Pigs. PLoS ONE, 13.","DOI":"10.1371\/journal.pone.0194385"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"429","DOI":"10.4103\/ijmr.ijmr_2737_21","article-title":"Development of Nipah Virus-Specific IgM & IgG ELISA for Screening Human Serum Samples","volume":"156","author":"Shete","year":"2022","journal-title":"Indian J. Med. Res."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"3000605251315913","DOI":"10.1177\/03000605251315913","article-title":"An Overview of ELISA: A Review and Update on Best Laboratory Practices for Quantifying Peptides and Proteins in Biological Fluids","volume":"53","author":"Aydin","year":"2025","journal-title":"J. Int. Med. Res."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"55","DOI":"10.1016\/j.jim.2008.10.002","article-title":"Validation and Comparison of Two Multiplex Technologies, Luminex and Mesoscale Discovery, for Human Cytokine Profiling","volume":"340","author":"Chowdhury","year":"2009","journal-title":"J. Immunol. Methods"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"1594","DOI":"10.3201\/eid1110.050513","article-title":"Genetic characterization of Nipah virus, Bangladesh, 2004","volume":"11","author":"Harcourt","year":"2005","journal-title":"Emerg. Infect. Dis."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"6690","DOI":"10.1128\/JVI.79.11.6690-6702.2005","article-title":"Receptor Binding, Fusion Inhibition, and Induction of Cross-Reactive Neutralizing Antibodies by a Soluble G Glycoprotein of Hendra Virus","volume":"79","author":"Bossart","year":"2005","journal-title":"J. Virol."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Monath, T.P., Nichols, R., Feldmann, F., Griffin, A., Haddock, E., Callison, J., Meade-White, K., Okumura, A., Lovaglio, J., and Hanley, P.W. (2023). Immunological Correlates of Protection Afforded by PHV02 Live Attenuated Recombinant Vesicular Stomatitis Virus Vector Vaccine against Nipah Virus Disease. Front. Immunol., 14.","DOI":"10.3389\/fimmu.2023.1216225"},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Alam, M., Rana, M.J., Salauddin, A., Bentley, E., Kamuyu, G., Shill, D.K., Jahan, S., Alam, M.M., Raihan, M.A., and Rahman, M.Z. (2025). Development and Validation of a Standardized Pseudotyped Virus-Based Neutralization Assay for Assessment of Anti-Nipah Virus Neutralizing Activity in Candidate Nipah Vaccines. Vaccines, 13.","DOI":"10.20944\/preprints202506.0472.v1"},{"key":"ref_15","unstructured":"Coalition for Epidemic Preparedness Innovations (2020, March 13). CEPI-Funded Nipah Virus Vaccine Candidate First to Reach Phase 1 Clinical Trial. Available online: https:\/\/cepi.net\/cepi-funded-nipah-virus-vaccine-candidate-first-reach-phase-1-clinical-trial."},{"key":"ref_16","first-page":"229","article-title":"Establishing an Immune Correlate of Protection for Nipah Virus in Nonhuman Primates","volume":"9","author":"Promeneur","year":"2024","journal-title":"npj Vaccines"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"May, A.J., and Acharya, P. (2024). Structural Studies of Henipavirus Glycoproteins. Viruses, 16.","DOI":"10.3390\/v16020195"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"406","DOI":"10.1002\/prca.201400130","article-title":"ELISA in the Multiplex Era: Potentials and Pitfalls","volume":"9","author":"Tighe","year":"2015","journal-title":"Proteom. Clin. Appl."},{"key":"ref_19","unstructured":"International Council for Harmonisation (2023). ICH Q2(R2): Validation of Analytical Procedures, ICH."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"151","DOI":"10.1186\/s12985-020-01425-8","article-title":"Effective Inactivation of Nipah Virus in Serum Samples for Safe Processing in Low-Containment Laboratories","volume":"17","author":"Watanabe","year":"2020","journal-title":"Virol. J."},{"key":"ref_21","unstructured":"(2023). WHO First International Standard for Nipah Virus Antibody (Standard No. WHO\/BS\/2023.2458)."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Satter, S.M., Aquib, W.R., Sultana, S., Sharif, A.R., Nazneen, A., Alam, M.R., Siddika, A., Ema, F.A., Chowdhury, K.I.A., and Alam, A.N. (2023). Tackling a Global Epidemic Threat: Nipah Surveillance in Bangladesh, 2006\u20132021. PLoS Neglected Trop. Dis., 17.","DOI":"10.1371\/journal.pntd.0011617"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"13581","DOI":"10.1038\/s41598-021-92826-4","article-title":"Validating Quantitative PCR Assays for Cell-Free DNA Detection without DNA Extraction in Exercising SLE Patients","volume":"11","author":"Neuberger","year":"2021","journal-title":"Sci. Rep."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Andreasson, U., Perret-Liaudet, A., van Waalwijk van Doorn, L.J., Blennow, K., Chiasserini, D., Engelborghs, S., Fladby, T., Genc, S., Kruse, N., and Kuiperij, H.B. (2015). A Practical Guide to Immunoassay Method Validation. Front. Neurol., 6.","DOI":"10.3389\/fneur.2015.00179"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"E260","DOI":"10.1208\/aapsj0902029","article-title":"Appropriate Calibration Curve Fitting in Ligand Binding Assays","volume":"9","author":"Findlay","year":"2007","journal-title":"AAPS J."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"50","DOI":"10.1016\/j.cca.2012.12.031","article-title":"Simultaneous liquid chromatography tandem mass spectrometric determination of urinary free metanephrines and catecholamines, with comparisons of free and deconjugated metabolites","volume":"418","author":"Peitzsch","year":"2013","journal-title":"Clin. Chim. Acta"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"851","DOI":"10.18609\/cgti.2025.098","article-title":"From Bench to Bedside: Navigating Bioanalytical Method Development and Validation for Clinical Efficacy and Regulatory Compliance","volume":"11","author":"Misra","year":"2025","journal-title":"Cell Gene Ther. Insights"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"228","DOI":"10.1097\/IM9.0000000000000190","article-title":"Indirect ELISAs Based on Nipah and Langya Virus Proteins for Detecting Antibodies in Animals","volume":"7","author":"Wu","year":"2025","journal-title":"Infect. Microbes Dis."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Zhu, W., Pickering, B., Smith, G., Pinette, M., Truong, T., Babiuk, S., Kobasa, D., Banadyga, L., and Yang, M. (2023). Development and Laboratory Evaluation of a Competitive ELISA for Serodiagnosis of Nipah and Hendra Virus Infection Using Recombinant Nipah Glycoproteins and a Monoclonal Antibody. Front. Vet. Sci., 10.","DOI":"10.3389\/fvets.2023.1120367"}],"container-title":["Vaccines"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-393X\/14\/6\/534\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,17]],"date-time":"2026-06-17T04:08:55Z","timestamp":1781669335000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-393X\/14\/6\/534"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,16]]},"references-count":29,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2026,6]]}},"alternative-id":["vaccines14060534"],"URL":"https:\/\/doi.org\/10.3390\/vaccines14060534","relation":{},"ISSN":["2076-393X"],"issn-type":[{"value":"2076-393X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,16]]}}}