Prakash Om, Goswami S V, Sehgal Neeta
Department of Zoology, University of Delhi, Delhi 110 007, India.
Comp Biochem Physiol C Toxicol Pharmacol. 2007 Nov;146(4):540-51. doi: 10.1016/j.cbpc.2007.06.008. Epub 2007 Jun 30.
Vitellogenin (Vg) and choriogenin (Chg) are sensitive biomarkers for testing endocrine disruption in fish. Therefore, we have developed immunoassays for Vg and Chg in the Indian freshwater murrel, Channa punctatus. Vg is a known precursor of egg-yolk proteins, whereas Chg contributes to the formation of egg-envelope. Vg and Chg were induced in male murrel by administration of estradiol-17beta. Chg had an apparent native molecular mass of 180 kDa. It consisted of a single peptide with a molecular mass of 110 kDa, whereas native Vg protein (530 kDa) contained 175 kDa peptide. Highly specific polyclonal antibodies against purified plasma proteins, Vg and Chg, were employed for developing competitive enzyme linked immunosorbent assays (ELISAs). The sensitivity of Vg assay was 3.9 ng/mL (working range 15-500 ng/mL) and of Chg assay was 1.56 ng/mL (working range 6-200 ng/mL). The inter- and intra-assay variations were well within acceptable limits. The two antisera did not cross-react with male plasma proteins. Antiserum to Vg did not cross-react with Chg. Similarly, antiserum to Chg showed no correlation with Vg. Further, immunofluorescence and Western blotting confirmed the specificity of Vg and Chg antisera.
卵黄蛋白原（Vg）和卵壳蛋白原（Chg）是用于检测鱼类内分泌干扰的敏感生物标志物。因此，我们开发了针对印度淡水鲃（Channa punctatus）中Vg和Chg的免疫测定方法。Vg是已知的蛋黄蛋白前体，而Chg有助于卵膜的形成。通过施用17β-雌二醇在雄性鲃中诱导出Vg和Chg。Chg的表观天然分子量为180 kDa。它由一个分子量为110 kDa的单一肽组成，而天然Vg蛋白（530 kDa）包含175 kDa的肽。针对纯化的血浆蛋白Vg和Chg的高度特异性多克隆抗体被用于开发竞争性酶联免疫吸附测定（ELISA）。Vg测定的灵敏度为3.9 ng/mL（工作范围15 - 500 ng/mL），Chg测定的灵敏度为1.56 ng/mL（工作范围6 - 200 ng/mL）。测定间和测定内的变异均在可接受范围内。两种抗血清与雄性血浆蛋白无交叉反应。Vg抗血清与Chg无交叉反应。同样，Chg抗血清与Vg无相关性。此外，免疫荧光和蛋白质印迹证实了Vg和Chg抗血清的特异性。
