{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,28]],"date-time":"2026-02-28T04:20:40Z","timestamp":1772252440413,"version":"3.50.1"},"reference-count":31,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2025,3,13]],"date-time":"2025-03-13T00:00:00Z","timestamp":1741824000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32473060"],"award-info":[{"award-number":["32473060"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32461120064"],"award-info":[{"award-number":["32461120064"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["2025A04J5445"],"award-info":[{"award-number":["2025A04J5445"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Guangzhou Basic and Applied Basic Research Project","award":["32473060"],"award-info":[{"award-number":["32473060"]}]},{"name":"Guangzhou Basic and Applied Basic Research Project","award":["32461120064"],"award-info":[{"award-number":["32461120064"]}]},{"name":"Guangzhou Basic and Applied Basic Research Project","award":["2025A04J5445"],"award-info":[{"award-number":["2025A04J5445"]}]},{"name":"Young Scholars of the Yangtze River Scholar Professor Program","award":["32473060"],"award-info":[{"award-number":["32473060"]}]},{"name":"Young Scholars of the Yangtze River Scholar Professor Program","award":["32461120064"],"award-info":[{"award-number":["32461120064"]}]},{"name":"Young Scholars of the Yangtze River Scholar Professor Program","award":["2025A04J5445"],"award-info":[{"award-number":["2025A04J5445"]}]},{"name":"Young Peal River Scholar of \u201cGuangdong Special Support Plan\u201d","award":["32473060"],"award-info":[{"award-number":["32473060"]}]},{"name":"Young Peal River Scholar of \u201cGuangdong Special Support Plan\u201d","award":["32461120064"],"award-info":[{"award-number":["32461120064"]}]},{"name":"Young Peal River Scholar of \u201cGuangdong Special Support Plan\u201d","award":["2025A04J5445"],"award-info":[{"award-number":["2025A04J5445"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>Interferon-\u03b3 (IFN-\u03b3), a member of the Type II IFN family, is a crucial cytokine in the immune system and serves as an important indicator of immune response. Intracellular cytokine staining (ICS) is a technique used to analyze the production of cytokines within individual cells, and it has a wide range of applications in the fields of immunological monitoring, vaccine trials, and the study of infectious diseases. This study aimed to prepare monoclonal antibodies against duck IFN-\u03b3 protein and to establish an ICS protocol for detecting the duck IFN-\u03b3 protein. The duIFN-\u03b3-His or duIFN-\u03b3-Fc gene was cloned into the pEE12.4 expression vector and expressed as a recombinant protein of size 20.2 KDa or 54.9 KDa in 293F cells. The purified recombinant proteins were inoculated into BALB\/c mice to generate splenic lymphocytes capable of secreting anti-duIFN-\u03b3 antibodies, and hybridoma cells were obtained after fusion with SP2\/0 cells. A new hybridoma cell line named 24H4, which stably secreted IgG3 \u03ba subtype antibody against duck IFN-\u03b3, was established. This monoclonal antibody (mAb) was identified by Western blot to recognize duck IFN-\u03b3 antibodies, and the indirect ELISA results showed that its ability to recognize IFN-\u03b3 protein reached 0.001 \u03bcg\/mL. The established ICS method was used to stain PBMCs after Concanavalin A (ConA) stimulation, and duck IFN-\u03b3 protein was successfully detected by flow cytometry, indicating that the ICS method was successful. In this study, we provide a crucial tool for subsequent research on duck cellular immune responses by using the monoclonal antibody 24H4.<\/jats:p>","DOI":"10.3390\/ani15060815","type":"journal-article","created":{"date-parts":[[2025,3,13]],"date-time":"2025-03-13T06:53:00Z","timestamp":1741848780000},"page":"815","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Development of a Monoclonal Antibody Against Duck IFN-\u03b3 Protein and the Application for Intracellular Cytokine Staining"],"prefix":"10.3390","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0009-0001-8736-0783","authenticated-orcid":false,"given":"Yingyi","family":"Chen","sequence":"first","affiliation":[{"name":"National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wei","family":"Song","sequence":"additional","affiliation":[{"name":"National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-6250-0938","authenticated-orcid":false,"given":"Junqiang","family":"Chen","sequence":"additional","affiliation":[{"name":"National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-1044-5793","authenticated-orcid":false,"given":"Chenyang","family":"Jin","sequence":"additional","affiliation":[{"name":"National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jiewei","family":"Lin","sequence":"additional","affiliation":[{"name":"National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ming","family":"Liao","sequence":"additional","affiliation":[{"name":"National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China"},{"name":"UK-China Centre of Excellence for Research on Avian Diseases, Guangzhou 510642, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6533-5456","authenticated-orcid":false,"given":"Manman","family":"Dai","sequence":"additional","affiliation":[{"name":"National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangdong Provincial Key Laboratory of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China"},{"name":"UK-China Centre of Excellence for Research on Avian Diseases, Guangzhou 510642, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,3,13]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1381","DOI":"10.1292\/jvms.11-0517","article-title":"Biological effects of chicken type III interferon on expression of interferon-stimulated genes in chickens: Comparison with type I and type II interferons","volume":"74","author":"Masuda","year":"2012","journal-title":"J. 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