{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,7]],"date-time":"2026-07-07T20:22:36Z","timestamp":1783455756264,"version":"3.55.0"},"reference-count":40,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2025,3,7]],"date-time":"2025-03-07T00:00:00Z","timestamp":1741305600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Vet. Sci."],"abstract":"<jats:sec><jats:title>Introduction<\/jats:title><jats:p><jats:italic>Listeria monocytogenes<\/jats:italic> causes zoonotic listeriosis with a high mortality rate, which is frequently detected in slaughterhouse processing environments and animal-based food. To enable the specific, rapid, and cost-effective detection of <jats:italic>L. monocytogenes<\/jats:italic> in environments and animal-based food, we developed a double-antibody sandwich quantitative ELISA (DAS-qELISA) method.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>The method is based on monoclonal antibodies targeting internalin G (InlG), a surface protein of <jats:italic>L. monocytogenes<\/jats:italic> with demonstrated immunogenicity. The antibody pair 1D2-2H10 was selected for use in the sandwich ELISA format. Optimization of the DAS-qELISA method was carried out to determine its detection limits for InlG protein and <jats:italic>L. monocytogenes<\/jats:italic>.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>The detection limits of the method were determined to be 32 ng\/mg for the InlG protein and 7875.83 CFU\/mL for <jats:italic>L. monocytogenes<\/jats:italic>. The accuracy of the method was evaluated across various bacterial concentrations, with results falling within 91.56\u2013107.07% and a coefficient of variation (CV) of less than 10%. Compared to traditional methods, this approach requires only 12 h of bacterial enrichment and incubation to achieve 100% accuracy.<\/jats:p><\/jats:sec><jats:sec><jats:title>Discussion<\/jats:title><jats:p>The DAS-qELISA developed in this study provides a rapid, accurate, and cost-effective tool for the detection of <jats:italic>L. monocytogenes<\/jats:italic> in environmental and animal-based food samples. This method could be a valuable addition to current diagnostic approaches, offering quicker turnaround times and high accuracy for pathogen detection.<\/jats:p><\/jats:sec>","DOI":"10.3389\/fvets.2025.1517845","type":"journal-article","created":{"date-parts":[[2025,3,7]],"date-time":"2025-03-07T10:58:46Z","timestamp":1741345126000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":7,"title":["Development of an internalin-based double-antibody sandwich quantitative ELISA for the detection of Listeria monocytogenes in slaughterhouse environments"],"prefix":"10.3389","volume":"12","author":[{"given":"Qing","family":"Cao","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenjing","family":"Shi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yanquan","family":"Wei","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiayu","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhonglong","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qian","family":"Chong","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qianqian","family":"Guo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kunzhong","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenyan","family":"Gai","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huitian","family":"Gou","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huiwen","family":"Xue","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2025,3,7]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"104776","DOI":"10.1016\/j.micpath.2021.104776","article-title":"Effect of Listeria monocytogenes on intestinal stem cells in the co-culture model of small intestinal organoids","volume":"153","author":"Huang","year":"2021","journal-title":"Microb Pathog"},{"key":"ref2","doi-asserted-by":"publisher","first-page":"584","DOI":"10.4269\/ajtmh.23-0029","article-title":"Placental listeriosis: case report and literature review","volume":"109","author":"Van der Merwe","year":"2023","journal-title":"Am J Trop Med Hyg"},{"key":"ref3","doi-asserted-by":"publisher","first-page":"1491","DOI":"10.3390\/pathogens11121491","article-title":"Listeria monocytogenes - how this pathogen uses its virulence mechanisms to infect the hosts","volume":"11","author":"Osek","year":"2022","journal-title":"Pathogens"},{"key":"ref4","doi-asserted-by":"publisher","first-page":"1447","DOI":"10.3390\/antibiotics11101447","article-title":"New insights into Listeria monocytogenes antimicrobial resistance, virulence attributes and their prospective correlation","volume":"11","author":"Elsayed","year":"2022","journal-title":"Antibiotics"},{"key":"ref5","doi-asserted-by":"publisher","first-page":"135","DOI":"10.3390\/microorganisms8010135","article-title":"Listeriosis outbreak in South Africa: a comparative analysis with previously reported cases worldwide","volume":"8","author":"Kaptchouang Tchatchouang","year":"2020","journal-title":"Microorganisms"},{"key":"ref6","doi-asserted-by":"publisher","first-page":"5500","DOI":"10.2903\/j.efsa.2018.5500","article-title":"The European Union summary report on trends and sources of zoonoses, zoonotic agents, and food-borne outbreaks in 2017","volume":"16","year":"2018","journal-title":"EFSA J"},{"key":"ref7","doi-asserted-by":"publisher","first-page":"459","DOI":"10.1089\/fpd.2017.2409","article-title":"The epidemiology of Listeria monocytogenes in China","volume":"15","author":"Li","year":"2018","journal-title":"Foodborne Pathog Dis"},{"key":"ref8","doi-asserted-by":"publisher","first-page":"1026","DOI":"10.3389\/fmicb.2015.01026","article-title":"Prevalence, enumeration, and pheno- and genotypic characteristics of Listeria monocytogenes isolated from raw foods in South China","volume":"6","author":"Chen","year":"2015","journal-title":"Front Microbiol"},{"key":"ref9","doi-asserted-by":"publisher","first-page":"92","DOI":"10.1016\/j.foodcont.2016.01.009","article-title":"Prevalence of food-borne pathogens in ready-to-eat meat products in seven different Chinese regions","volume":"65","author":"Yang","year":"2016","journal-title":"Food Control"},{"key":"ref10","doi-asserted-by":"publisher","first-page":"1074","DOI":"10.5740\/jaoacint.11-311","article-title":"Evaluation of applied biosystems MicroSEQ real-time PCR system for detection of Listeria spp. in food and environmental samples","volume":"95","author":"Petrauskene","year":"2012","journal-title":"J AOAC Int"},{"key":"ref11","doi-asserted-by":"publisher","first-page":"110475","DOI":"10.1016\/j.ijfoodmicro.2023.110475","article-title":"A novel multiplex PCR-based method for the detection of Listeria monocytogenes clonal complex 8","volume":"409","author":"Cheng","year":"2024","journal-title":"Int J Food Microbiol"},{"key":"ref12","doi-asserted-by":"publisher","first-page":"37","DOI":"10.1016\/j.mimet.2015.08.012","article-title":"Quantification of Listeria monocytogenes cells with digital PCR and their biofilm cells with real-time PCR","volume":"118","author":"Klan\u010dnik","year":"2015","journal-title":"J Microbiol Methods"},{"key":"ref13","doi-asserted-by":"publisher","first-page":"349","DOI":"10.4014\/jmb.2308.08043","article-title":"Detection of Listeria species by conventional culture-dependent and alternative rapid detection methods in retail ready-to-eat foods in Turkey","volume":"34","author":"Dincer","year":"2024","journal-title":"J Microbiol Biotechnol"},{"key":"ref14","doi-asserted-by":"publisher","first-page":"186","DOI":"10.1089\/fpd.2022.0079","article-title":"Targeting novel LPXTG surface proteins with monoclonal antibodies for immunomagnetic separation of Listeria monocytogenes","volume":"20","author":"Zhang","year":"2023","journal-title":"Foodborne Pathog Dis"},{"key":"ref15","doi-asserted-by":"publisher","first-page":"340085","DOI":"10.1016\/j.aca.2022.340085","article-title":"Sandwich fluorometric method for dual-role recognition of Listeria monocytogenes based on antibiotic-affinity strategy and fluorescence quenching effect","volume":"1221","author":"Li","year":"2022","journal-title":"Anal Chim Acta"},{"key":"ref16","doi-asserted-by":"publisher","first-page":"1407","DOI":"10.1111\/mmi.14836","article-title":"Role of internalin proteins in the pathogenesis of Listeria monocytogenes","volume":"116","author":"Ireton","year":"2021","journal-title":"Mol Microbiol"},{"key":"ref17","doi-asserted-by":"publisher","first-page":"123","DOI":"10.1007\/s13197-022-05597-9","article-title":"Combined antibodies against internalins a and B proteins have potential application in immunoassay for detection of Listeria monocytogenes","volume":"60","author":"Lopes-Luz","year":"2023","journal-title":"J Food Sci Technol"},{"key":"ref18","doi-asserted-by":"publisher","first-page":"1328","DOI":"10.1099\/mic.0.067744-0","article-title":"Occurrence of mutations impairing sigma factor B (SigB) function upon inactivation of Listeria monocytogenes genes encoding surface proteins","volume":"159","author":"Quereda","year":"2013","journal-title":"Microbiology"},{"key":"ref19","doi-asserted-by":"publisher","first-page":"222","DOI":"10.1089\/fpd.2006.0081","article-title":"Distribution of internalin gene profiles of Listeria monocytogenes isolates from different sources associated with phylogenetic lineages","volume":"4","author":"Jia","year":"2007","journal-title":"Foodborne Pathog Dis"},{"key":"ref20","doi-asserted-by":"publisher","first-page":"63","DOI":"10.33073\/pjm-2022-009","article-title":"The characteristics and function of internalin G in Listeria monocytogenes","volume":"71","author":"Gou","year":"2022","journal-title":"Pol J Microbiol"},{"key":"ref21","doi-asserted-by":"publisher","first-page":"4069","DOI":"10.11843\/j.issn.0366-6964.2024.09.031","article-title":"Preparation of monoclonal antibodies against internalin G of Listeria monocytogenes and their preliminary application","volume":"55","author":"Ma","year":"2024","journal-title":"Chin J Anim Vet Sci"},{"key":"ref22","doi-asserted-by":"publisher","first-page":"2666","DOI":"10.1128\/aem.57.9.2666-2670.1991","article-title":"Phosphatidylinositol-specific phospholipase C activity as a marker to distinguish between pathogenic and nonpathogenic Listeria species","volume":"57","author":"Notermans","year":"1991","journal-title":"Appl Environ Microbiol"},{"key":"ref23","doi-asserted-by":"publisher","first-page":"1412","DOI":"10.1271\/bbb.70252","article-title":"Species-specific detection of Listeria monocytogenes using polymerase chain reaction assays targeting the prfA virulence gene cluster","volume":"73","author":"Jung","year":"2009","journal-title":"Biosci Biotechnol Biochem"},{"key":"ref24","doi-asserted-by":"publisher","first-page":"769","DOI":"10.3390\/foods12040769","article-title":"The prevalence and antibiotic-resistant of Listeria monocytogenes in livestock and poultry meat in China and the EU from 2001 to 2022: a systematic review and meta-analysis","volume":"12","author":"Zhang","year":"2023","journal-title":"Food Secur"},{"key":"ref25","doi-asserted-by":"publisher","first-page":"356","DOI":"10.1080\/22221751.2021.1888658","article-title":"Prevalence and transmission characteristics of Listeria species from ruminants in farm and slaughtering environments in China","volume":"10","author":"Zhao","year":"2021","journal-title":"Emerg Microbes Infect"},{"key":"ref26","doi-asserted-by":"publisher","first-page":"2307520","DOI":"10.1080\/22221751.2024.2307520","article-title":"Risk factors associated with food consumption and food-handling habits for sporadic listeriosis: a case-control study in China from 2013 to 2022","volume":"13","author":"Niu","year":"2024","journal-title":"Emerg Microbes Infect"},{"key":"ref27","doi-asserted-by":"publisher","first-page":"52","DOI":"10.1016\/j.cofs.2020.09.002","article-title":"Advances in typing and identification of foodborne pathogens","volume":"37","author":"Wei","year":"2021","journal-title":"Curr Opin Food Sci"},{"key":"ref28","doi-asserted-by":"publisher","first-page":"297","DOI":"10.4014\/jmb.1310.10013","article-title":"Advances in rapid detection methods for foodborne pathogens","volume":"24","author":"Zhao","year":"2014","journal-title":"J Microbiol Biotechnol"},{"key":"ref29","doi-asserted-by":"publisher","first-page":"1111","DOI":"10.3390\/microorganisms11051111","article-title":"A review of modern methods for the detection of foodborne pathogens","volume":"11","author":"Aladhadh","year":"2023","journal-title":"Microorganisms"},{"key":"ref30","doi-asserted-by":"publisher","first-page":"167","DOI":"10.1186\/s12951-022-01376-y","article-title":"Development of nanobody-horseradish peroxidase-based sandwich ELISA to detect Salmonella Enteritidis in milk and in vivo colonization in chicken","volume":"20","author":"Gu","year":"2022","journal-title":"J Nanobiotechnology"},{"key":"ref31","doi-asserted-by":"publisher","first-page":"7025","DOI":"10.3168\/jds.2016-11320","article-title":"Sensitive detection of Escherichia coli O157:H7 based on cascade signal amplification in ELISA","volume":"99","author":"Shan","year":"2016","journal-title":"J Dairy Sci"},{"key":"ref32","doi-asserted-by":"publisher","first-page":"8325","DOI":"10.1128\/AEM.06507-11","article-title":"Ruminant rhombencephalitis-associated Listeria monocytogenes alleles linked to a multilocus variable-number tandem-repeat analysis complex","volume":"77","author":"Balandyt\u00e9","year":"2011","journal-title":"Appl Environ Microbiol"},{"key":"ref33","doi-asserted-by":"publisher","first-page":"137","DOI":"10.47276\/lr.78.2.137","article-title":"Serum samples from patients with mycobacterial infections cross-react with HIV structural proteins Gp41, p55, and p18","volume":"78","author":"Hussain","year":"2007","journal-title":"Lepr Rev"},{"key":"ref34","doi-asserted-by":"publisher","first-page":"422","DOI":"10.1016\/S0006-291X(03)01193-8","article-title":"Homology modeling of allergenic cyclophilins: IgE-binding site and structural basis of cross-reactivity","volume":"307","author":"Roy","year":"2003","journal-title":"Biochem Biophys Res Commun"},{"key":"ref35","doi-asserted-by":"publisher","first-page":"S1","DOI":"10.1096\/fasebj.26.1_supplement.lb200","article-title":"Evaluation of immunoassay interference reduction technologies","volume":"26","author":"Jentz","year":"2012","journal-title":"FASEB J"},{"key":"ref36","doi-asserted-by":"publisher","first-page":"8310","DOI":"10.1021\/acs.analchem.9b01077","article-title":"Simply mixing poly protein G with detection antibodies enhances the detection limit and sensitivity of immunoassays","volume":"91","author":"Chen","year":"2019","journal-title":"Anal Chem"},{"key":"ref37","doi-asserted-by":"publisher","first-page":"531","DOI":"10.3109\/00365513.2012.702352","article-title":"Reduction of non-specific binding in immunoassays requiring long incubations","volume":"72","author":"Farajollahi","year":"2012","journal-title":"Scand J Clin Lab Invest"},{"key":"ref38","doi-asserted-by":"publisher","first-page":"4","DOI":"10.1002\/bdrb.21047","article-title":"Analytic evaluation of a human ELISA kit for measurement of inhibin B in rat samples","volume":"98","author":"Coulson","year":"2013","journal-title":"Birth Defects Res B Dev Reprod Toxicol"},{"key":"ref39","doi-asserted-by":"publisher","first-page":"691","DOI":"10.5713\/ajas.14.0868","article-title":"Quality evaluation of five commercial enzyme-linked immunosorbent assay kits for detecting aflatoxin B1 in feedstuffs","volume":"28","author":"Sun","year":"2015","journal-title":"Asian Australas J Anim Sci"},{"key":"ref40","doi-asserted-by":"publisher","first-page":"156","DOI":"10.1016\/j.foodcont.2017.03.042","article-title":"A sandwich-type ELISA for the detection of Listeria monocytogenes using the well-oriented single chain Fv antibody fragment","volume":"79","author":"Liu","year":"2017","journal-title":"Food Control"}],"container-title":["Frontiers in Veterinary Science"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fvets.2025.1517845\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,3,7]],"date-time":"2025-03-07T10:58:49Z","timestamp":1741345129000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fvets.2025.1517845\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,3,7]]},"references-count":40,"alternative-id":["10.3389\/fvets.2025.1517845"],"URL":"https:\/\/doi.org\/10.3389\/fvets.2025.1517845","relation":{},"ISSN":["2297-1769"],"issn-type":[{"value":"2297-1769","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,3,7]]},"article-number":"1517845"}}