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Serological diagnosis is a cost\u2010effective approach for disease surveillance, but a major drawback of current serological tests is the cross\u2010reactivity with other bacteria that results in false positive reactions. Development of indirect tests with improved sensitivity and specificity that use selected <jats:italic>B. canis<\/jats:italic> proteins instead of the whole antigen remain a priority.<\/jats:p><\/jats:sec><jats:sec><jats:title>Experimental Design<\/jats:title><jats:p>A western blotting assay was developed to define the serum antibody patterns associated to infection using a panel of positive and negative dog sera. <jats:italic>B. canis<\/jats:italic> positive sera recognized immunogenic bands ranging from 7 to 30\u00a0kDa that were then submitted to ESI\u2013LC\u2010MS\/MS and analyzed by bioinformatics tools.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>A total of 398 <jats:italic>B. canis<\/jats:italic> proteins were identified. Bioinformatics tools identified 16 non cytoplasmic immunogenic proteins predicted as non\u2010homologous with the most important <jats:italic>Brucella<\/jats:italic> cross\u2010reactive bacteria and nine <jats:italic>B. canis<\/jats:italic> proteins non\u2010homologous to <jats:italic>B. ovis<\/jats:italic>; among the latter, one resulted non\u2010homologous to <jats:italic>B. melitensis<\/jats:italic>. Data are available via ProteomeXchange with identifier PXD042682.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions and Clinical Relevance<\/jats:title><jats:p>The western blotting test developed was able to distinguish between infected and non\u2010infected animals and may serve as a confirmatory test for the serological diagnosis of <jats:italic>B. canis<\/jats:italic>. The mass spectrometry and <jats:italic>in silico<\/jats:italic> results lead to the identification of specific candidate antigens that pave the way for the development of more accurate indirect diagnostic tests.<\/jats:p><\/jats:sec>","DOI":"10.1002\/prca.202200116","type":"journal-article","created":{"date-parts":[[2023,8,3]],"date-time":"2023-08-03T01:45:53Z","timestamp":1691027153000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Proteomics and bioinformatics investigations to improve serological diagnosis of canine brucellosis"],"prefix":"10.1002","volume":"17","author":[{"given":"Mirella","family":"Luciani","sequence":"first","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ivanka","family":"Krasteva","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0181-8184","authenticated-orcid":false,"given":"Tiziana","family":"Di Febo","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fabrizia","family":"Perletta","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Federica","family":"D'Onofrio","sequence":"additional","affiliation":[{"name":"Faculty of Bioscience and Technology for Food Agriculture and Environment University of Teramo Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fabrizio","family":"De Massis","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nicola","family":"D'Alterio","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Flavio","family":"Sacchini","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Manuela","family":"Tittarelli","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise Teramo Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2023,8,2]]},"reference":[{"key":"e_1_2_9_2_1","first-page":"355","volume-title":"Manual of diagnostic tests and vaccines for terrestrial animals","author":"W. 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