Follicle-Stimulating Hormone (FSH) Immunoactivity in Porcine Follicular Fluid Is Not Pituitary FSH*

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Abstract
Two inhibitors of FSH binding to receptor have been isolated from porcine follicular fluid and shown to have in vitro biological activity. These inhibitors were distinct separable entities with opposite biological effects (agonist and antagonist) on cultured FSH-response Sertoli cells. In light of the fact that the agonist-containing fraction (P4) inhibited [125I]human (h) FSH binding to anti-FSH antiserum as well as to receptor, characterization of this factor was undertaken to determine its relationship to pituitary FSH. The P4 fraction was further purified by affinity chromatography, which removed a major protein from immunoreactive components. Western blotting of sodium dodecyl sulfate-polyacrylamide gels using polyclonal (anti-hFSH.beta.) antibodies revealed a major immunoreactive band at 55,000 mol wt (Mr). When electrophoresed under reducing conditions, major immunoreactive proteins at 58,000 and 45,000 Mr were identified. These bands were also observed in extracts from bovine testes and raw porcine follicular fluid after electrophoresis and Western blotting. Whereas the monoclonal antibody used to characterize this inhibitor does not recognize porcine pituitary FSH, the Mr of the immunoreactive proteins are greater than that of pituitary FSH, and the immunoreactive bands do not reduce to subunits, as observed for pituitary FSH under reducing conditions, we concluded that gonadal extracts contain FSH-immunoreactive proteins that are immunologically and biochemically distinguishable from pituitary FSH. While the physiological role of these proteins remains to be determined, their presence in gonadal extracts or fluids vitiates assessment of FSH within the gonad by RIA using antiserum against hFSH.