{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,3]],"date-time":"2026-02-03T18:09:18Z","timestamp":1770142158672,"version":"3.49.0"},"reference-count":21,"publisher":"Wiley","issue":"3","license":[{"start":{"date-parts":[[2024,3,4]],"date-time":"2024-03-04T00:00:00Z","timestamp":1709510400000},"content-version":"vor","delay-in-days":3,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Current Protocols"],"published-print":{"date-parts":[[2024,3]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>The main challenge in the \u201cpost\u2010GWAS\u201d era is to determine the functional meaning of genetic variants and their contribution to disease pathogenesis. Development of suitable mouse models is critical because disease susceptibility is triggered by complex interactions between genetic, epigenetic, and environmental factors that cannot be modeled by in vitro models. Thyroglobulin (TG) is a key gene for autoimmune thyroid disease (AITD) and several single nucleotide polymorphisms (SNPs) in the TG coding region have been associated with AITD. The classical model of experimental autoimmune thyroiditis (EAT), based on immunization of genetically susceptible mouse strains with purified TG protein in adjuvant, does not allow testing the impact of TG sequence variants on the development of autoimmune thyroiditis. Here we describe a protocol for the induction of EAT by immunization of mice susceptible to thyroiditis with an adenovirus vector carrying full\u2010length human TG cDNA (Ad\u2010TG EAT). We also provide support protocols for evaluation of autoimmune thyroiditis including serological assessment of TG antibodies, in vitro splenocyte proliferation assay and cytokines secretion, thyroid histology, and evaluation of thyroid lymphocytic infiltration by immunostaining. This protocol for EAT induction allows manipulation of the TG cDNA to introduce variants associated with AITD, enabling the testing of the functional effects of susceptible variants and their haplotypes on the immunogenicity of TG. Furthermore, the Ad\u2010TG EAT mouse model is a valuable model for studying the interactions of the TG variants with non\u2010genetic factors influencing AITD development (e.g., cytokines, iodine exposure) or with variants of other susceptible genes (e.g., HLA\u2010DR\u03b21). \u00a9 2024 Wiley Periodicals LLC.<\/jats:p><jats:p><jats:bold>Basic Protocol<\/jats:bold>: Development of a mouse model of autoimmune thyroiditis induced by immunization with adenovirus containing full\u2010length thyroglobulin cDNA<\/jats:p><jats:p><jats:bold>Support Protocol 1<\/jats:bold>: Splenocytes isolation<\/jats:p><jats:p><jats:bold>Support Protocol 2<\/jats:bold>: T cell stimulation and carboxyfluorescein diacetate succinimidyl ester (CFSE) based cell proliferation assay<\/jats:p><jats:p><jats:bold>Support Protocol 3<\/jats:bold>: Cytokine assays: measuring levels of interferon gamma (IFN\u03b3) and interleukins IL\u20102, IL\u20104, and IL\u201010 in splenocyte supernatants<\/jats:p><jats:p><jats:bold>Support Protocol 4<\/jats:bold>: Evaluating thyroid histology and infiltration with immune cells: hematoxylin\u2010eosin staining of mice thyroid glands<\/jats:p><jats:p><jats:bold>Support Protocol 5<\/jats:bold>: Immunohistochemistry of thyroid tissues: Immunofluorescence protocol of paraffin\u2010embedded thyroid sections<\/jats:p><jats:p><jats:bold>Support Protocol 6<\/jats:bold>: Anti\u2010thyroglobulin antibody measurement in mice sera by enzyme\u2010linked immunosorbent assay (ELISA)<\/jats:p>","DOI":"10.1002\/cpz1.938","type":"journal-article","created":{"date-parts":[[2024,3,4]],"date-time":"2024-03-04T10:34:33Z","timestamp":1709548473000},"source":"Crossref","is-referenced-by-count":1,"title":["A Flexible Mouse Model of Autoimmune Thyroiditis Induced by Immunization with an Adenovirus Containing Full\u2010Length Thyroglobulin cDNA"],"prefix":"10.1002","volume":"4","author":[{"given":"Aizhan","family":"Kozhakhmetova","sequence":"first","affiliation":[{"name":"Division of Endocrinology, Department of Medicine Albert Einstein College of Medicine  Bronx New York"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yaron","family":"Tomer","sequence":"additional","affiliation":[{"name":"Division of Endocrinology, Department of Medicine Albert Einstein College of Medicine  Bronx New York"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1590-2547","authenticated-orcid":false,"given":"Mihaela","family":"Stefan\u2010Lifshitz","sequence":"additional","affiliation":[{"name":"Division of Endocrinology, Department of Medicine Albert Einstein College of Medicine  Bronx New 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