IgG抗体适体与16kD抗原联合检测血清结核抗体研究

The research of IgG antibodies aptamers and 16 kD antigen in TB serological diagnosis

目的研究结核患者IgG抗体适体与16 kD抗原组合在结核病血清学检测中的应用价值。方法 SELEX技术筛选得到结核患者IgG抗体适体,并将适体构建的混合ELISA体系用于少量血清标本的检测,初步鉴定适体的检测效果,然后将适体与16 kD抗原联合构建ELISA检测体系进行221份血清标本的分析。结果 SELEX筛选得到4个高亲和适体;单适体血清学检测结果显示,适体之间存在交叉互补性;适体两两组合检测效果显示,N1+N6组合的阳性平均值与阴性平均值的比值最高;221份血清标本检测结果显示,N1+N6适体组合、16 kD抗原以及N1+N6+16 kD组合3种检测体系的敏感性和特异性分别为47.31%和92.97%、49.46%和91.41%、51.61%和92.19%,N1+N6适体组合、16 kD抗原以及N1+N6+16 kD组合3种检测体系的结核病组和健康对照组之间的t检验结果依次为t=6.564,P<0.0001;t=5.279,P<0.0001;t=4.348,P<0.0001,结核病组和非结核疾病对照组之间的t检验结果依次为t=4.196,P<0.0001;t=3.014,P=0.0031;t=2.929,P=0.004,差异均具有统计学意义,三种检测方法之间卡方检验显示,差异无统计学意义(P>0.05);单独适体组合与单独抗原检测合并计算的敏感性、特异性为67.74%、84.38%,与N1+N6+16 kD组合检测结果的卡方检验显示,敏感性差异有统计学意义(χ2=8.52,P<0.05),特异性差异无统计学意义(χ2=2.04,P>0.05)。结论 SELEX技术筛选获得的特异性IgG抗体适体和16KD抗原联合具有良好的结核病血清学诊断价值,可用于结核病的实验室诊断。

Objective To explore the potential laboratory diagnosis value of IgG antibodies aptamers and 16 kD antigen in TB(tuberculosis) serological combination diagnosis.Methods Aptamers against TB IgG antibodies were obtained by SELEX technology.ELISA systems based on the aptamers and/or 16 kD antigen were used to evaluate the potential diagnosis value of these methods by detecting clinical serum samples.Results Four aptamers with higher affinity were obtained and combined with each other to build mixed-sandwich ELISA methods.8 cases of clinical serum samples from TB and healthy control were tested by these methods.Detection systems based on N1 and N6 aptamer combinations,which had the highest ratio of average A value of TB group to control group,were selected and further evaluated by 221 cases of clinical serum samples.ELISA systems based on 16 kD antigen or combinations of aptamers and antigen were also evaluated by these cases.The differences of TB group and healthy group were statistically significant(P<0.05) in all these three detection systems,and the differences of TB group and non-TB control group were also statistically significant(P<0.05).The sensitivity and the specificity was 47.31% and 92.97%(based on N1 and N6 aptamers),49.46% and 91.41%(based on 16 kD antigen),51.61% and 92.19%(based on the combinations of aptamers and antigen).The detection effects showed no statistical differences among these three detection systems(P>0.05).Otherwise,the sensitivity and specificity consolidated from independent detection systems based on aptamers or 16 kD antigen was 67.74% and 84.38%.The sensitivity showed statistical differences(χ2=8.52,P<0.05),while the specificity not(χ2=2.04,P>0.05).Conclusions The ELISA methods based on aptamers and 16 kD protein will play an important role in the serological diagnosis of tuberculosis.