Immunocytochemical demonstration of chicken hypophyseal thyrotropes and development of a radioimmunological indicator for chicken TSH using monoclonal and polyclonal homologous antibodies in a subtractive strategy.
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The production and characterization of monoclonal antibodies (mAbs) toward the chicken pituitary glycoproteins are described. MAbs were first subdivided on an immunocytochemical (ICC) basis: one category of mAbs reacts with both thyrotropic and gonadotropic cells whereas another stains exclusively gonadotropic cells. The hypothesis that the former are anti-glycoprotein alpha and that the latter are anti-beta LH and/or anti-beta FSH mAbs was further elaborated in a homologous tracer binding assay. The putative anti-beta gonadotropin mAbs exclusively recognize iodinated USDA-cLH-I-1, whereas the supposed anti-alpha mAbs react with both USDA-cLH-I-1 and USDA-cFSH-I-1. The ICC localization of pituitary thyrotropes has been achieved in a double-staining protocol using an anti-LH mAb to subtract specifically the gonadotropic cells from the complete alpha-subunit-containing cell population. The result is a strictly cephalic cell population that is densely granulated in hyperthyroid and hardly detectable in hypothyroid birds. A similar subtractive approach was used to develop a thyroid-stimulating hormone (TSH)-indicator in RIA. The total titer of alpha-containing molecules is first quantified with an anti-alpha mAb. The contribution of both LH and FSH to this titer is then measured by homologous USDA-RIA and subtracted from the total alpha-immunoreactivity to give an indication of the TSH level of the sample. The direct effect of thyroid-releasing hormone on pituitary TSH release in vitro in juvenile chicks and the effect of hypo- and hyperthyroidism on circulating TSH levels in vivo in juvenile broilers have been demonstrated.