Quantitative determination of the gonadotrope polypeptide (GP87) contained in and release by gonadotrope cells using a specific anti-rat GP87

A polyclonel antibody has been reised against the gonadotrope polypeptide (GP87), a secretogranin II form released by rat gonadotrope cells under LHRH stimulation. GP87 was previously isolated through bidirectional polyacrylamide gel electrophoresis followed by electrotransfer onto nitrocellulose membrane. Using this antibody, it could be shown by immunoelectron microscopy that GP87 was localised in the secretory granules of several restricted cell types. A specific quantitative immunoassay was then devised to determine the cell content of GP87 or its release into culture medium. This assay involved an SDS-polyacrylamide gel electrophoresis of proteins followed by western blotting with anti-GP87 and densitometry of the nitrocellulose membrane. Combining this assay with preliminary separation of cell populations by centrifugal elutriation, it was demonstrated that GP87 is highly concentrated in gonadotropes. Moreover, the LHRH-stimulated minute-to-minute release of GP87 by rat pituitary cell aggregates in perifusion xas shown to be closely correlated to that of LH especially in its biphasic pattern. This anti-rat GP87 thus appears highly suitable to analyse the intracel1ular compartmentation ofsecretogranin II/GP87 and putative modification processes preceding or following LHRH-induced exocytosis