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Localized immune response in dermal leishmaniasis results in low systemic antibody production. Consequently, serologic diagnostic platform should focus on detecting\n                    <jats:italic>Leishmania<\/jats:italic>\n                    antigens directly from the skin lesions. Designing multiepitope\u2010based proteins as an alternative to crude antigens can enhance sensitivity and specificity, lower production costs, and facilitate development of more field\u2010amenable diagnostic tools. A chimeric multiepitope protein construct (THAG) was designed from four\n                    <jats:italic>Leishmania<\/jats:italic>\n                    spp. genes (\n                    <jats:italic>txnpx<\/jats:italic>\n                    ,\n                    <jats:italic>gp63<\/jats:italic>\n                    ,\n                    <jats:italic>hsp70<\/jats:italic>\n                    , and\n                    <jats:italic>amastin<\/jats:italic>\n                    ). Bioinformatic tools were employed to ensure that the construct was optimized for expression in\n                    <jats:italic>Escherichia coli<\/jats:italic>\n                    and had favorable physicochemical properties. The gene was synthesized, subcloned into the pET28a(+) vector, and expressed in\n                    <jats:italic>E. coli<\/jats:italic>\n                    BL21 (DE3). Protein purification was carried out using Ni\u2010NTA affinity chromatography under hybrid conditions. The purified protein was used to generate polyclonal antibodies in rabbits, and the immunoreactivity of antiserum was assessed against\n                    <jats:italic>Leishmania<\/jats:italic>\n                    antigens from lesions of 54 volunteers with suspected active skin lesions mimic CL through an in\u2010house ELISA. The THAG multiepitope recombinant protein of\n                    <jats:italic>Leishmania<\/jats:italic>\n                    has successfully designed, expressed, purified and confirmed in our study. The results demonstrated sensitivity, specificity and accuracy of the ELISA test in detection of\n                    <jats:italic>Leishmania<\/jats:italic>\n                    antigens in CL lesions to be 90.91% (75.67%\u201398.08%), 95.24% (76.18%\u201399.88%), and 92.59% (82.11%\u201397.94%), respectively. This multiepitope protein has the potential to be further evaluated as a diagnostic tool for CL patients. While we have evaluated this multiepitope protein in an ELISA format, it might be applied to other test formats especially in immunochromatographic assays for the rapid detection of\u00a0\n                    <jats:italic>Leishmania<\/jats:italic>\n                    antigens in skin lesions. Furthermore, it has the potential to be used as a tool to detect anti\u2010\n                    <jats:italic>Leishmania<\/jats:italic>\n                    antibodies in serum samples of canine and human visceral leishmaniasis (VL) patients.\n                  <\/jats:p>","DOI":"10.1002\/bab.70159","type":"journal-article","created":{"date-parts":[[2026,3,29]],"date-time":"2026-03-29T18:25:29Z","timestamp":1774808729000},"page":"2135-2148","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Development of a Recombinant Chimeric Antigen Expressing Different Epitopes of\n                    <i>Leishmania<\/i>\n                    spp. For Diagnosis of Human Cutaneous Leishmaniasis"],"prefix":"10.1002","volume":"73","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-4317-7271","authenticated-orcid":false,"given":"Mahmoud","family":"Nateghi\u2010Rostami","sequence":"first","affiliation":[{"name":"Department of 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