{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,9]],"date-time":"2026-04-09T18:37:05Z","timestamp":1775759825633,"version":"3.50.1"},"reference-count":42,"publisher":"Public Library of Science (PLoS)","issue":"4","license":[{"start":{"date-parts":[[2026,4,9]],"date-time":"2026-04-09T00:00:00Z","timestamp":1775692800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001695","name":"Japan Science and Technology Corporation","doi-asserted-by":"publisher","award":["JPMJSP2122"],"award-info":[{"award-number":["JPMJSP2122"]}],"id":[{"id":"10.13039\/501100001695","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>The bovine genetic variant A2 beta-casein is associated with fewer digestive and absorption issues compared to A1 beta-casein, leading to increased global demand for A2 milk. However, contamination with the A1 variant during collection, transportation, or sterilization of A2 milk poses a risk, necessitating a verification test to ensure A2 milk does not contain A1 beta-casein. We developed an A1-specific monoclonal antibody (mAb) and a general mAb that reacts with both A1 and A2 variants, using the iliac lymph node method. A sandwich ELISA was created using the general mAb as the capture antibody and the A1-specific mAb as the detection antibody to identify A1 beta-casein in milk. This ELISA successfully detected A1 beta-casein in raw and pasteurized A2 milk, including ultra-high temperature treated milk. The test identified A1 beta-casein when the A1 spike in A2 milk exceeded 1% in volume, indicating its capability to detect contamination from one A1A1 cow in a herd of one hundred A2A2 cows. The developed A1 beta-casein ELISA is suitable for high-throughput analysis and valuable for monitoring A1 beta-casein contamination in commercially produced A2 milk.<\/jats:p>","DOI":"10.1371\/journal.pone.0345548","type":"journal-article","created":{"date-parts":[[2026,4,9]],"date-time":"2026-04-09T17:40:55Z","timestamp":1775756455000},"page":"e0345548","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["Development of a new sandwich ELISA for the detection of bovine A1 beta-casein"],"prefix":"10.1371","volume":"21","author":[{"given":"Ayumi","family":"Watanabe","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tomoe","family":"Kobayashi","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Anna","family":"Okamoto","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Daiki","family":"Oka","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tomohiro","family":"Noguchi","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ren","family":"Ozawa","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Koumei","family":"Shirasuna","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2606-059X","authenticated-orcid":true,"given":"Makoto","family":"Matsuyama","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9441-4922","authenticated-orcid":true,"given":"Takashi","family":"Kuramoto","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"340","published-online":{"date-parts":[[2026,4,9]]},"reference":[{"issue":"2","key":"pone.0345548.ref001","doi-asserted-by":"crossref","first-page":"252","DOI":"10.3390\/ani10020252","article-title":"Frequencies Evaluation of \u03b2-Casein Gene Polymorphisms in Dairy Cows Reared in Central Italy","volume":"10","author":"C Sebastiani","year":"2020","journal-title":"Animals (Basel)"},{"issue":"8","key":"pone.0345548.ref002","doi-asserted-by":"crossref","first-page":"957","DOI":"10.1016\/S0196-9781(99)00088-1","article-title":"Enzymatic release of neocasomorphin and beta-casomorphin from bovine beta-casein","volume":"20","author":"Y Jinsmaa","year":"1999","journal-title":"Peptides"},{"issue":"1","key":"pone.0345548.ref003","doi-asserted-by":"crossref","first-page":"82","DOI":"10.1186\/s12937-016-0201-x","article-title":"Clinical evaluation of glutathione concentrations after consumption of milk containing different subtypes of \u03b2-casein: results from a randomized, cross-over clinical trial","volume":"15","author":"R Deth","year":"2016","journal-title":"Nutr J"},{"issue":"2","key":"pone.0345548.ref004","doi-asserted-by":"crossref","first-page":"141","DOI":"10.3390\/molecules21020141","article-title":"Detecting \u03b2-Casein Variation in Bovine Milk","volume":"21","author":"AM Caroli","year":"2016","journal-title":"Molecules"},{"issue":"24","key":"pone.0345548.ref005","doi-asserted-by":"crossref","first-page":"15637","DOI":"10.3390\/ijms232415637","article-title":"Does a Little Difference Make a Big Difference? 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