Nagler J J, Hwang S J, Idler D R
Marine Sciences Research Laboratory, Memorial University of Newfoundland, St. John's, Canada.
Gen Comp Endocrinol. 1991 Dec;84(3):374-80. doi: 10.1016/0016-6480(91)90084-j.
Growth hormone (GH) was isolated from sockeye salmon (Oncorhynchus nerka) pituitary glands using established techniques of affinity and gel filtration chromatography, and preparative polyacrylamide gel electrophoresis. The GH activity was followed throughout the fractionation procedure with a bioassay based on the increase of serum triiodothyronine (T3) in rainbow trout (Oncorhynchus mykiss), measured by radioimmunoassay (RIA). Amino-terminal amino acid sequence analysis and subsequent comparison with established GH sequences from other Oncorhynchus sp. were used to confirm the isolation of sockeye salmon GH (ssGH). The bioassay was sensitive to a dose of 55 ng of purified ssGH/g fish. Monomeric GHs, located in the carbohydrate-poor protein fraction, were the only pituitary components that elevated serum T3. Twenty-four hours after GH injection was an appropriate and practical time to blood sample, allowing completion of the bioassay, including RIA, in 3 days. The generic homology, between the source of pituitaries and the bioassay animals used in this study, should permit the bioassay to be useful during GH isolation from pituitaries of all Oncorhynchus sp.
利用亲和层析、凝胶过滤层析和制备聚丙烯酰胺凝胶电泳等成熟技术，从红大马哈鱼（Oncorhynchus nerka）的垂体中分离出生长激素（GH）。在整个分级分离过程中，通过基于放射免疫分析（RIA）测定虹鳟鱼（Oncorhynchus mykiss）血清三碘甲状腺原氨酸（T3）增加的生物测定法来跟踪GH活性。通过氨基末端氨基酸序列分析以及随后与其他大麻哈鱼属物种已确定的GH序列进行比较，来确认红大马哈鱼GH（ssGH）的分离。该生物测定法对55 ng纯化的ssGH/g鱼的剂量敏感。位于低糖蛋白组分中的单体GH是唯一能提高血清T3的垂体成分。注射GH后24小时是采集血样的合适且实用的时间，这使得在3天内能够完成包括RIA在内的生物测定。本研究中垂体来源与所用生物测定动物之间的类属同源性，应使该生物测定法在从所有大麻哈鱼属物种的垂体中分离GH时有用。
