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Commercially available IgG-ELISA kits cannot distinguish between anti-\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      IgG in inactivated bacterin-induced hyperimmune sera and convalescent sera resulting from natural\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      infection. Establishment of an ELISA to detect anti-\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      IgG in convalescent sera will facilitate the evaluation of the\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      status of pig farms.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      In this study, we expressed and purified recombinant Mhp366-N protein, which contains an epitope recognized by\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      convalescent sera but not hyperimmune sera, for use as a coating antigen. For the\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      convalescent serum IgG-ELISA, the optimal antigen concentration, blocking buffer, blocking time, dilution of serum, incubation time with serum, secondary antibody dilution, secondary antibody incubation time and colorimetric reaction time were 0.25\u00a0\u00b5g\/mL, 2.5\u2009% skim milk, 1\u00a0h, 1:500, 0.5\u00a0h, 1:10,000, 1\u00a0h and 15\u00a0min, respectively. Validation of the\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      convalescent serum IgG-ELISA showed a cut-off value of 0.323, the intra-assay CV ranged from 3.27 to 7.26\u2009%, the inter-assay CV ranged from 3.46 to 5.93\u2009%, and the assay was able to differentiate convalescent sera from antibodies to 7 other porcine respiratory pathogens. The convalescent serum IgG-ELISA detected no anti-\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      IgG in hyperimmune serum samples while a commercial IgG-ELISA identified 95\/145 of these sera as positive. The accuracy of the\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      convalescent serum IgG-ELISA was comparable to the sIgA-ELISA but better than the commercial IgG-ELISA.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>\n                      The convalescent serum IgG-ELISA is a reproducible, sensitive, and specific indirect ELISA to detect anti-\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      IgG in naturally infected pathogen-induced convalescent sera. This ELISA could be used to carry out large scale surveillance of\n                      <jats:italic>M. hyopneumoniae<\/jats:italic>\n                      infection in pig farms regardless of vaccination status.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1186\/s12917-021-02828-7","type":"journal-article","created":{"date-parts":[[2021,4,9]],"date-time":"2021-04-09T08:33:54Z","timestamp":1617957234000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["Development of an indirect ELISA for detection of anti-Mycoplasma hyopneumoniae IgG in naturally infected pathogen-induced convalescent sera"],"prefix":"10.1186","volume":"17","author":[{"given":"Yaqin","family":"Tian","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zuobo","family":"Xu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yukang","family":"Wen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mei","family":"Yang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yaru","family":"Ning","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhaodi","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6978-0277","authenticated-orcid":false,"given":"Honglei","family":"Ding","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2021,3,16]]},"reference":[{"key":"2828_CR1","doi-asserted-by":"crossref","unstructured":"Maes D, Sibila M, Kuhnert P, Segal\u00e9s J, Haesebrouck F, Pieters M. 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All experimental protocols were approved by the Institutional Animal Ethics Committee of Southwest University (Approval no. IACUC-20170921-3) and performed accordingly. The objectives, protocols and potential risks were clearly explained to all participating farm owners. Written informed consent was obtained from all participating farm owners.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethics approval and consent to participate"}},{"value":"Not applicable.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Consent for publication"}},{"value":"The authors declare that they have no competing interests.","order":4,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"123"}}