1995 Fiscal Year Final Research Report Summary
Immunobiochemical study on growth hormone and vitellogenin in salmonids
| Project/Area Number |
06660253
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Fisheries chemistry
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| Research Institution | HOKKAIDO UNIVERSITY |
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Principal Investigator |
HARA Akihiko Hokkaido Univ., Fac.of Fish., Prof., 水産学部, 教授 (40091483)
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| Project Period (FY) |
1994 – 1995
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| Keywords | salmonids / growth hormone / vitellogenin / ELISA |
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| Research Abstract |
Growth hormone (GH) was purified from the pituitary glands of chum salmon (Oncorhynchus keta). The purification of GH was performed by saltingout, ion exchange chromatography and gel filtration. The purified protein was identified as chum salmon GH by SDS-PAGE and Western blotting using the antiserum obtained from a rabbit immunized with recombinant chum salmon GH (a-rsGH).
A specific and sensitive enzyme-linked immunosorbent assay (ELISA) was developed for the measurement of low levels of serum GH in chum salmon. The non-competitive ELISA was performed by a sandwich method using polyclonal a-GH rabbit IgG as the first antibody, its biotinylated Fab' fragment as the second antibody, and the avidin-biotin reaction for signal amplification. This assay could be run in 3 days and routinely detected GH at concentrations as low as 0.5 ng/ml. The development of an ELISA for GH made it possible to quantify of serum GH levels. This ELISA cross-reacted with pituitary extracts and sera from four salmonid genera (Oncorhynchus, Salmo, Salvelinus and Hucho). Moreover, dilution curves of purified chum salmon GH and sera from several species of the genus Oncorhynchus were parallel to the standard curve.
During May, approximately four months prior to spawning, serum GH levels increased, reaching peak levels in Masu salmon (O.masou). At this time body weight also increased. By June, levels of GH declined and relatively stable until September. From June to September, body weight in the fish also remained constant. Serum vitellogenin (Vg) levels increased during May, reaching peak levels in August and then declined during September. Interestingly, the increase in serum Vg levels followed the increase in serum GH which occurred during May.
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