长白山林蛙输卵管糖蛋白多抗制备及应用研究

Preparation and Application of Polyclonal Antibodies against Oviduct Glucoprotein in Rana Chensinensis Changbaishanensis

针对长白山林蛙输卵管糖蛋白制备特异性多克隆抗体。将新鲜林蛙输卵管蛋白粗提液经SepharoseFastFlow阴离子交换柱和SephadexG-100凝胶层析柱获得纯化的输卵管糖蛋白组分。以纯化输卵管糖蛋白免疫新西兰大白兔,制备多抗血清,采用辛酸-饱和硫酸铵法对多抗进行纯化并鉴定。结果表明:纯化后的多抗ELISA效价为1:64000,纯化多抗具有较高的敏感性和特异性,效价较高。利用制备的多抗对长白山林蛙输卵管分泌细胞进行免疫组织化学定位,为分泌细胞培养产物中输卵管糖蛋白进行检测提供研究条件。

In order to prepare specific polyclonal antibodies against oviduct glucoprotein （OGP） in Rana Chensinensis Changbaishanensis, the crude fresh OGP-rich extract was purified by Sepharose fast flow anion exchange column and Sephadex G-100 gel column chromatography, and New-Zealand albino rabbits were immunized with the purified OGP for the generation of polyclonal antisera. The purification and identification of the polyclonal antisera obtained were performed using the caprylic acid-saturated ammonium sulfate method. The titer of the purified polyclonal antibodies was measured by enzyme linked immunosorbent assay to be 1：64000, which were characterized by high sensitivity, specificity and titer. Moreover, the polyclonal antibodies prepared in this study were used for the immunohistochemical localization and detection of OGP secreting cells in the oviduct of Rana Chensinensis Changbaishanensis, which was expected to lay an experimental foundation for determining OGP in the culture product of OGP secreting cells.