蝙蝠血清IgG的纯化及兔抗蝙蝠IgG酶标抗体的制备

目的纯化蝙蝠血清IgG,制备兔抗蝙蝠IgG酶标抗体。方法采用亲和层析纯化法纯化蝙蝠血清IgG,SDS-PAGE电泳鉴定蝙蝠IgG纯度。免疫大白兔制备兔抗蝙蝠IgG抗血清,免疫双扩散法测定抗血清效价,亲和层析纯化法纯化抗血清IgG。用改良过碘酸钠标记法制备兔抗蝙蝠IgG酶标抗体,直接ELISA和Western blot法对兔抗蝙蝠IgG酶标抗体进行工作浓度测定。结果纯化的蝙蝠血清IgG,其SDS-PAGE测定纯度大于95%;免疫大白兔所制备的抗血清免疫双扩散效价为1∶64;用改良过碘酸钠标记法制备兔抗蝙蝠IgG酶标抗体,其直接ELISA和Western blot工作浓度分别为1∶12800和大于1∶2000。结论制备了蝙蝠血清IgG的抗血清和酶标抗体,为蝙蝠的血清学检测体系提供了技术和资源储备。

Purification of IgG from Bat＇s Serum and the Production of Rabbit Anti-bat IgG-HRP

Objective To purify IgG from Bat＇s serum and prepare the rabbit anti-bat IgG-HRP. Methods The IgG from bat＇ s serum was purified using Hitrap Protein-A Affinity Chromatography and the purity of the bat-IgG was identified by SDS-PAGE analysis. The rabbit anti-bat IgG from the antiserum was purified by affinity chromatography and the IgG was labeled with HRP by the modified sodium periodate method. The working concentration of the rabbit anti-bat IgG-HRP was detected by direct ELISA and Western blotting. Results The purified bat IgG was obtained and the purity was up to 95% on SDS-PAGE. The titer of the rabbit antisrum was up to 1 ： 64 on Two-Dimensional immune Diffusion. The working concentration of the HRP-rabbit anti-bat IgG was up to 1 ： 12800 for direct ELISA and 1 ： 2000 for Western blotting. Conclusion The purified IgG of bat＇ s serum and the rabbit anti-bat polyclonal IgO-HRP was obtained. It would be a useful tool to develope the serological test system for bat.