兔抗阴道毛滴虫重组蛋白AP65多克隆抗体的制备、纯化及鉴定

Preparation,purification and identification of rabbit polyclonal antibody against recombinant protein AP65 of Trichomonas vaginalis

目的制备、纯化并鉴定兔抗阴道毛滴虫重组蛋白AP65的多克隆抗体。方法日本大耳兔多次免疫重组蛋白AP65,间接ELISA法检测抗体效价,用饱和硫酸铵法和n protein A sepharose 4FF柱纯化多克隆抗体,Western blot检测免疫反应性。结果第3次加强免疫后,间接ELISA法检测血清效价达到1∶25600,用硫酸铵法纯化多克隆抗体纯度为56%,而后采用亲合层析柱n protein A sepharose 4FF再次纯化,纯度达到80%。Western blot鉴定兔抗重组蛋白AP65多克隆抗体能识别重组蛋白AP65。结论成功制备了高效价、高纯度的抗阴道毛滴虫重组蛋白AP65多克隆抗体。

Objective To prepare, purify and identify the rabbit polyelonal antibody against fusion AP65 of Trichomonas vaginalis. Methods The fusion AP65 was used to immunize rabbits for the antibody. The titer of antise-rum was demonstrated by indirect EI.ISA. The polyclonal antibodies were purified by saturation ammonium sulfate and n protein A sepharose 4 FF. The immunogenicity of the ployclonal antibody was determined by western blot. Results The titers of antibody detected by indirection ELISA reached to 1 ; 25 600. The purification of the polyclonal antibody was highly 80% by n protein A sepharose 4 FF after purified by ammonium sulfate, which was 56% however. The specification was confirmed by western blot. Conclusion The high purity and high titer polyclonal antibody was prepared successfully.