剑尾鱼卵黄蛋白原的ELISA检测

以卵黄脂磷蛋白(lipovitellin，Lv)抗血清为抗体，以纯化的卵黄蛋白原(vitellogenin，Vtg)为抗原，建立了间接酶联免疫吸附反应(ELISA)方法检测剑尾鱼(Xiphophorus helleri)雄鱼整体匀浆液中ρ(Vtg).结果表明，该方法的检测灵敏度为7.8 ng/mL，批内变异系数为5.094%，批间变异系数为5.540%，工作范围为32.5～2 000 ng/mL，在该范围内，标准曲线具有良好的线性和重复性.由于该方法可直接在1块或在不同的酶标板上准确地进行比较，因而可利用ELISA方法测定经诱导雄鱼整体匀浆液中ρ(Vtg)水平.结果显示，50 μg/L 17-β雌二醇(E₂)诱导21 d的雄性剑尾鱼有明显的Vtg产生；10 μg/L E₂诱导剑尾鱼亦有Vtg产生，但较50 μg/L E₂诱导组低；1 μg/L E₂诱导组及对照组剑尾鱼没有Vtg产生，检测孔OD450/对照OD₄₅₀(P/N)值小于2.1. 

Development of ELISA for Detecting Xiphophorus helleri Vitellogenin

An indirect competitive enzyme-linked immunosorbent assay (ELISA) was established for detecting vitellogenin (Vtg) in the whole body homogenate (WBH) of male swordtail fish (Xiphophorus helleri). This technique was developed with antiserum of lipovitellin(Lv)resistance as antibody and Vtg as antigen. The resultsindicated that the working range was 32.5～2 000 ng/mL; the sensitivity was 7.8 ng/mL; intra-assay and inter-assay coefficients of variation were 5.094% and 5.540% respectively. The technique shows good linearity and repeatability in standard curve. WBH of male swordtail fish sample can be checked inone same coated well and different coated wells to develop screening of environmental estrogen. The results also indicated that the WBH from 50 μg/L 17-β estradiol treated with 21 days male had Vtg obviously, and that of 10 μg/L had too, but that of 1 μg/L had not yet (P/N was less than 2.1).