利用罗非鱼卵黄原蛋白建立雌激素污染的检测方法

Establishment of estrogens detection method using Nile tilapia(Oreochromis niloticus) vitellogenin

中文关键词:  环境雌激素  卵黄原蛋白  罗非鱼  多克隆抗体

英文关键词:environmental estrogens  vitellogenin  Oreochromis niloticus  polyclonal antibody

中文摘要:

      目的 以尼罗罗非鱼为实验对象, 以卵黄原蛋白(Vg)为生物标志物开发环境雌激素的生物检测技术。方法 采用凝胶过滤与离子交换层析相结合的方法从17β-雌二醇诱导后的罗非鱼血浆中分离纯化Vg, 对纯化的蛋白进行鉴定后制备多克隆抗血清, 建立Vg的酶联免疫吸附测定实验(ELISA), 并用于样品测定。结果 建立的ELISA工作范围为15.6~1000 ng/mL, 组内与组间差异分别为6.85%与6.79%, 能有效检测100、500、1000 μg/L双酚A对雄鱼血浆Vg的诱导效应。结论 本研究建立的Vg ELISA具有较高的敏感度、特异性与精确度, 可为水体环境雌激素污染检测提供重要工具。

英文摘要:

      Objective To develop a bioassay method for monitoring environmental estrogens with Nile tilapia vitellogenin (Vg) as biomarker. Methods Vg was purified from the 17β-estradiol induced plasma of Nile tilapia by gel filtration and anion-exchange chromatography, identified and made into polyclonal antibody. Then, a sensitive enzyme-linked immunosorbent assay (ELISA) was developed to quantify Vg. Results The established ELISA had a working range from 15.6 to 1000 ng/mL, and the intra- and inter-assay coefficients of variations were 6.85% and 6.79%, respectively. The ELISA could effectively detect the estrogenic activity of bisphenol A. Conclusion The highly sensitive, specific and robust Nile tilapia Vg ELISA method is suitable for the detection of water environmental estrogens.