新型α-半乳糖苷酶多克隆抗体的制备及特异性鉴定

Preparation and Identification of Polyclonal Antibody Against a Novel α-Galactosidase from Bacteroides fragilis

目的:制备高效价、高特异性的新型α-半乳糖苷酶的兔多抗,并鉴定该抗体的特异性。方法:用脆弱类杆菌来源的基因重组α-半乳糖苷酶(纯度大于90%)免疫新西兰大白兔,获得α-半乳糖苷酶的兔抗血清,并经HiTrap rProtein A柱纯化获得高纯度的抗体;用间接ELISA法检测抗体效价,Western印迹评价抗体的特异性。结果:通过免疫法得到了α-半乳糖苷酶的兔多克隆抗体血清,抗体效价达1∶1×106,经rProtein A柱纯化后获得了高效价、高纯度的抗体,Western印迹显示该抗体特异性地与新型α-半乳糖苷酶结合。结论:获得了新型α-半乳糖苷酶的高效价、高特异性的兔多克隆抗体,可用于血型转变过程中残留α-半乳糖苷酶含量的特异性检测。

Objective： To prepare high titer and high specific rabbit polyclonal antibody against a novel α-galactosidase from Bacteroides fragilis. Methods： The New Zealand rabbits were immunized with purified recombinant bacteria α-galactosidase（the purity〉90%）. Rabbit sera were purified by HiTrap rProtein A column, and its titer and specificity were detected by ELISA and Western blotting respectively Results： The purity of antibody protein was about 95%. And the titer of rabbit sera were 1：1 ~ 106. Western blotting showed that the antibody reacted with α-galactosidase only. Conclusion： We had obtained high titer and high purity rabbit polyclonal antibody of α-galactosidase. The antibody can be used in detecting the minimal amount of residual α-galactosidase involved in conversion of blood type B to O.