人巨细胞病毒pp65抗体检测方法的建立

Establishment of an ELISA test for detecting anti-human cytomegalovirus pp65 IgG antibody

目的建立检测人巨细胞病毒pp65IgG抗体（HCMV pp65 IgG）的间接酶联免疫吸附试验（ELISA）。方法通过棋盘滴定法选择包被抗原和酶标二抗的最适浓度：以商品抗HCMV pp65单抗为阳性模拟标本，建立检测抗HCMV pp65 IgG抗体，并对临床收集的各类血清标本进行检测和分析。结果检测间接ELISA HCMV pp65 IgG的最佳包被抗原浓度为10ng／孔，酶标二抗的工作浓度为1：3000。当阳性吸光度似）值为0．298时，87份不同组的血清标本（HCMV感染活动组、HCMV感染不活动组、HCMV未感染组）阳件检出率分别为51．6％（16／31）、12．1％（4／33）和0．0％（0／23）。3组两两间差异均存在显著性（P〈0．01）。结论本研究建立了检测HCMV pp65抗体的方法，应用该方法能从相应人群中检出该抗体，检测不存在假阳性，并发现该抗体与抗HCMV IgM的检出有关。

Objective To establish a new enzyme-linked immunosorbent assay（ELISA） for detection of anti-human cytomegalovirus（HCMV） pp65 IgG in serUm. Methods Screening the best condition of the amount of pp65 protein coated and the dilution of horseradish peroxidase （HRP） labeled anti-human IgG for this test. The commercial anti-HCMV pp65 IgG was used as positive specimen. Analysis of different groups of sertan samples were tested by the established method. Results The best condition for this ELISA test was that the coating amount of pp65 protein was 10 ng per well, the best dilution of HRP labeled anti-human IgG was 1：3 000, the minimum d etectiori of the simulant positive specimen Was 1：5.12× 10^4.When the cut-off value of this method was A=0.29g, among 87 sera of 3 groups （active HCMV infection, Nonactive HCMV infection, without HCMV infection） were tested, the positive results of each group were 51.6% （16/31）,12.1% （4/33） and 0.0% （0/23） respectively. There was significant difference between each two groups （P〈0.01）. Conclusions The established ELISA method could detect anti-HCMV pp65 IgG antibody, there was no false positive result, and the HCMV pp65 IgG antibody was related with anti-HCMV IgM antibodies.