Immunoidentification of gonadotropin releasing hormone receptor in human sperm, pituitary and cancer cells

Abstract

Problem: To generate monoclonal antibodies specific to the receptor of gonadotropin releasing hormone (GnRH) for immunoidentification of the GnRH receptor (RGnRH) in human sperm, pituitary and cancer cells.

Methods: Four oligopeptides corresponding to RGnRH amino acid residues 1-29, 182-193, 195-206 and 293-306 in the extracellular domains were synthesized, conjugated to hemocyanin from Keyhole Limpet and used as immunogens to generate specific monoclonal antibodies. Enzyme-linked immunosorbent assay was used for initial screening of hybridomas.

Results: A total of 15 hybrid cell lines secreting RGnRH-specific monoclonal antibodies were initially established. By using some of these monoclonal antibodies as immunohistochemical probes, RGnRH was localized in the acrosomal region of human sperm, in the anterior pituitary tissue and in cancer cells of human ovarian and cervical origins. RGnRH was shown to have a molecular size of 45,000 +/- 5,000 kDa by Western blot assay. Expression of RGnRH mRNA in several human tissues and cancer cells was established by the reverse transcriptase-polymerase chain reaction method.

Conclusion: RGnRH-specific monoclonal antibodies may be a valuable tool of identifying the presence of RGnRH in various normal and malignant human tissues.