Purification of chinook salmon (Oncorhynchus tshawytscha) GH for receptor study

A method for the purification of chinook Salmon (Oncorhynchus tshawytscha) GH, which retains its biological activity, is described. The biological activity was investigated with an established radioreceptor assay using liver membranes from pregnant rabbits and bovine GH as standard and labelled hormone. The enrichment of the preparation was checked with electrophoresis (SDS-PAGE). Extraction and further steps were carried out using low molarity alkaline buffer (pH 8–10, M = 100 mM). Three chromatography steps were performed (Concanavalin-A sepharose, Bio-gel P60, DEAE). Ion exchange chromatography was performed under isocratic conditions (using a 50 cm column). Two isoforms (sGH1 and sGH2) were isolated. The purification yield is 0.7% compared to lyophilized pituitaries. The molecule is homogeneous in SDS-PAGE. Contamination by prolactin, gonadotrophin and corticotrophin is negligible (< 0.5%). It could be demonstrated that the biological activity of the preparation is maintained since this preparation stimulates the growth of juvenile trout (Salmo gairdneri) and binds specifically (35%) to trout liver membranes.     

多种动物血清中生物活性LH/CG的放射受体测定法(RRA)

利用绵羊卵巢黄体细胞膜作为 LH / CG受体来源 ,以放射性12 5 I- h CG作为示踪标记配体 ,以高纯度的h CG为标准品建立标准曲线 ,建立了一种有效的 LH- RRA测定方法。它能够直接检测多种动物血清中与受体结合的具有生物活性的 LH的动态变化。该法测定程序简单 ,在 2 h内完成并获得可重复性结果。测定方法的灵敏度为 0 .3ng/管 ,批内和批间误差分别为 ( 3.4 1± 3.14 ) %和 ( 8.70± 7.93) %。动物 (牛、兔、猪 )垂体液中 LH含量反应曲线和 h CG标准曲线平行良好。牛、羊、猪、兔、鼠和仓鼠血清中 LH的剂量反应曲线和h CG标准曲线的平行性表明 ,该 LH- RRA可以用来检测不同动物血清中的具有生物活性的 LH变化。回收试验表明 ,在该测定方法中加入 3、10、30 ng h CG时的回收率分别为 ( 10 0 .59± 5.55) %、( 99.55± 9.86 ) %、( 10 4 .2 8± 7.4 5) %,说明测定结果的正确性。该 LH - RRA的测定结果反映了动物正常的生理状态。本试验垂体中有活性的 LH水平远远高于外周循环中的水平 ,雌、雄动物循环血液中的 LH含量不一致。     

牦牛促黄体素放射受体分析方法的建立

以大鼠睾丸组织匀浆作受体,~(125)I—hCG作放射配体,hCG作参考标准,且在标准曲线各测定管中加入与待测血浆等量的无LH牦牛血浆,籍以排除血浆干扰因子的影响,成功地建立了LH/hCG的放射受体分析方法。试验表明,该法是一种测定牦牛LH的灵敏可靠的方法。