抗霍乱弧菌O139型的单克隆抗体免疫印迹分析抗原表位及应用

Recognition of Anti-VC O139 McAbs to Antigen Epitopes by Western Blotting

目的 制备霍乱弧菌O139型（Vibrio cholerae O139，VC O139）单克隆抗体后应用免疫印迹技术识别抗原表位特性。方法 SDS—PAGE结合Western blotting以及APAAP方法。结果 霍乱弧菌0139型抗原经还原剂处理，SDS—PAGE11％-20％梯度胶电泳，转印染色后，能分辨出20余条清晰的蛋白带和脂多糖片状图谱。3株抗霍乱弧茵0139型单抗的抗原表位，主要集中于20～70kD相对分子质量的蛋白带，其中0139的2株单抗O139—14、O139—15分别与68—50kD、45—25kD、20～14kD对应VC O139的蛋白质抗原表位起反应，而另外一株单抗，0139—92只与65kD、20kD的抗原表位特异性结合。结论 VC0139的0139—14、O139—15McAb同对应抗原表位点的结合分布较广，有利于对VC O139抗原的快速、敏感、特异检测。

Objective To prepare the anti -VC O139 McAbs and investigate the recognition of antigen epitopes by Western blotting. Methods Combining SDS - PAGE method with Western blotting method and APAAP method. Results Being treated by reducing agents, the antigens of VC O139 could be distinguished for more than 10 protein zones by gradient gel SDS - PAGE 11% - 20%. These zones were then transferred and stained by HRP （ horseradish peroxidase） and a color of brown was shown with light background. The re- suits showed that the 2 McAbs of O139 - 14, O139 - 15 against VC O139 reacted with protein antigens of 68 -50 kD,45 -25 kD,20 -14 kD, respectively, and the another McAb of O139 -92 against VC O139, reacted with protein antigens of 65 kD,20 kD respectively. Conclusions McAbs of O139 - 14, O139 - 15 against VCO139 could combined with different antigenic determinates in a wide range, thus, which can make the detection of the VC O139 antigens to be rapid, sensitive and specific.