血清p53抗体ELISA检测方法的建立

Development of ELISA method for detecting serum p53 antibodies

目的建立一种检测血清p53抗体的间接ELISA方法。方法以重组p53蛋白作为包被抗原，检测ELISA方法的敏感性、特异性、精密度与稳定性，同时对203份正常人血清及548份临床确诊恶性肿瘤病人血清进行检测。结果p53蛋白的最佳包被浓度为1μg／mL。包被液为0．05mol／L的碳酸盐缓冲液。3份p53抗体阳性血清稀释至1：400时，仍均为阳性；重复8次其变异系数范围为3．67％-16．88％。中和阻断试验结果表明阻断率随着p53蛋白含量的增加而增高。203份健康人血清的p53抗体全部为阴性，548份病人血清有78例阳性，阳性率为14．23％。结论该方法敏感、特异、稳定，可用于人群血清p53抗体的检测。

Objective To develop a specific indirect ELISA method for detecting anti - p53 antibodies in serum. Methods The recombinant p53 protein was used as the coating antigen to test sensitivi- ty, specificity, precision and stability of ELISA method. 203 serum samples from healthy individuals and 548 from patients with malign tumor were examined. Results The optimal concentration of the recombinant protein was 1μg/mL and the coating buffer was 0. 05 mol/L carbonic acid buffer （ pH9. 5 ）. 3 sera remained positive when they were diluted by the ration of 1 ： 400, and the variation coefficient was 3.67% - 16. 88% after repeatedly assaying for 8 times. Blocking test of specific IgG showed the rate of blocking increased with the p53 protein. Serum samples from healthy persons observed no positive （0/203） whereas 14. 23% （78/548） of patients with tumor were positive. Conclusion The new developed ELISA method for p53 antibodies detection was wide range, fine sensitivity, high precision and stability.