两种不同示踪剂用于蛋白质与抗原分子微阵列信号检测的对比研究  

Comparative investigation of signal detection for protein and antigen microarray by using two different tracers

作　　者：虞伟[1] 顾宁[1] 武建国[2] 葛存旺[1] 廖建辉[2] 

机构地区：[1]东南大学分子与生物分子电子学教育部重点实验室,南京210096 [2]南京军区南京总医院全军医学检验中心临床免疫实验室,南京210002

出　　处：《东南大学学报（自然科学版）》2003年第1期64-67,共4页Journal of Southeast University：Natural Science Edition

基　　金：国家自然科学基金资助项目 ( 6 0 1710 0 5)

摘　　要：以氧化型琼脂糖凝胶膜为基片 ,制作蛋白质与抗原分子微阵列 ,建立了检测固相人IgG、游离IgG和自身抗体分子的免疫学测定模式 ,分别以辣根过氧化物酶二氨基联苯胺双氧水 (HRP DAB H2 O2 )和胶体金硝酸银对苯二酚 2种示踪系统作为蛋白质与抗原分子微阵列信号显示剂 ,并对其作用的原理与特点进行讨论 .检测结果发现基于胶体金的免疫金银染色法 (IGSS)比固相酶免疫测定法(EIA)敏感 .2种示踪系统对固相人IgG的最低检出量分别为 0 .0 5ng和 0 .5ng;而对血清游离IgG的最低检出量IGSS法比EIA法至少高出 1 0倍 .两法用于自身抗体的测定亦表现出较好的重复性 ,在 1 4例HCV感染者中 ,自身抗体的总体检出率分别为 3 8.6%和 3 1 .4% ,结果具有显著的同步性 .故同时辅以 2种示踪剂并用于阵列芯片的联合测试 。By using oxidiated agarose-gel membrane as a matrix, protein and antigen microarrays were prepared and subsequent several immunoassays were developed for solid-phase and sample free IgG as well as autoantibodies. In this assay, two display-systems, horseradish peroxidase-diaminobenzidine tetrahydrochloride-hydrogen peroxide(HRP-DAB-H2O2)and colloidal gold-AgNO3-hydroquinone, were utilized as signal detection of microarrays, and also their interaction principles and characteristic which displayed were discussed. The results show that the detective sensitivity of immunogold-silver staining(IGSS) is higher than that of solid-phase enzyme immunoassay(EIA). The minimum solid-phase human IgG limits are 0.05 ng and 0.5 ng respectively. Whereas the lowest serum IgG detected by IGSS is at least 10 times higher than that of EIA. The autoantibodies also show that a good reproducibility is achieved by both methods. The positive rate of autoantibodies in 14 cases with HCV infection are 38.6% (IGSS) and 31.4% (EIA) respectively. The results acquired by using these tracer systems are significantly consistent. Thus the levels of autoantibodies can be improved by using two tracers for microarray detection simultaneously.

关 键 词：示踪剂 抗原分子微阵列 信号检测 对比研究 酶免疫测定法 免疫金银染色法 蛋白质 示踪系统 

分 类 号：Q503[生物学—生物化学] R446.6[医药卫生—诊断学]