title: Radioimmunoassay of relaxin-like gonad-stimulating peptide in the starfish Patiria (=Asterina) pectinifera.
abstract: A relaxin-like gonad-stimulating peptide (RGP) from starfish Patiria (=Asterina) pectinifera is the first identified invertebrate gonadotropin for final gamete maturation. An antiserum against P. pectinifera RGP (PpeRGP) was produced by immunizing rabbits with a PpeRGP sulfanyl-polyethylene glycol derivative conjugated with keyhole limpet hemocyanin (KLH) as the antigen. The antiserum was used for the development of a specific and sensitive radioimmunoassay (RIA) for the measurement of RGP. In binding experiments using radioiodinated PpeRGP and antiserum against PpeRGP, a displacement curve was obtained using radioinert PpeRGP. The sensitivity of the RIA, defined as the amount of PpeRGP that significantly decreased the counts by 2 SD from the 100% bound point, averaged 0.040±0.002pmol PpeRGP per 100μl assay buffer (0.40±0.02nM) in 10 assays. Intra-assay and inter-assay coefficients of variation were 6.1% and 2.7%, respectively. Serial dilution of whole homogenates from the radial nerve cords and circumoral nerve-rings of P. pectinifera produced displacement curves parallel to the PpeRGP standard. Thus, the amounts of PpeRGP were determined as 1.54±0.09pmol/mg wet weight of radial nerves and 0.87±0.04pmol/mg wet weight of nerve-rings, respectively. On contrary, pyloric stomach, pyloric caeca, tube-feet, ovaries, testes, and ovarian follicle cells did not react in the RIA system. Furthermore, the A- and B-chains of PpeRGP, Asterias amurensis RGP, bovine insulin, and human relaxin did not show cross-reactivity in the RIA. These results strongly suggest that the RIA system is a highly specific and sensitive with respect to PpeRGP.
authors.affiliation: Department of Biology, Faculty of Education and Integrated Sciences, Center for Advanced Biomedical Sciences, Waseda University, Wakamatsucho 2-2, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Kazutoshi
authors.initials: K
authors.lastname: Yamamoto
authors.affiliation: Tateyama Marine Laboratory, Marine and Coastal Research Center, Ochanomizu University, Kou-yatsu 11, Tateyama, Chiba 294-0301, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Kiyomoto
authors.affiliation: Department of Applied Biochemistry, School of Engineering, Tokai University, Kitakaname 4-1-1, Hiratsuka, Kanagawa 259-1292, Japan.
authors.forename: Hidekazu
authors.initials: H
authors.lastname: Katayama
authors.affiliation: Department of Biology, Faculty of Education, Tokyo Gakugei University, Nukuikita-machi 4-1-1, Koganei-shi, Tokyo 184-8501, Japan. Electronic address: bio-mita@u-gakugei.ac.jp.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Mita
mesh_terms: D000818:Animals; D047288:Asterina / Q000254:growth & development / Q000378:metabolism*; D006066:Gonads / Q000378:metabolism*; D007447:Invertebrate Hormones / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011863:Radioimmunoassay / Q000379:methods*; D012065:Relaxin / Q000378:metabolism*
chemical_list: D007447:Invertebrate Hormones; D010446:Peptide Fragments; D012065:Relaxin
keywords: Quantitative method; Radial nerve cords; Radioimmunoassay; Relaxin-like gonad-stimulating peptide; Starfish gonadotropin