同时检测4种传染病病原体的蛋白质微阵列研究

A Protein Micro-array for Simultaneous Detection of 4 Pathogens for Infectcon Diseases

〔目的〕建立1种用蛋白质微阵列法可同时检测血清中艾滋病病毒(HIV)抗体、丙型肝炎病毒(HCV)抗体、梅毒螺旋体(TP)抗体和乙型肝炎病毒表面抗原的方法。〔方法〕将基因工程HIV、HCV、TP重组抗原和乙肝病毒单克隆抗体共价结合于固相载体玻片上,制成蛋白质微阵列。血清样本经稀释、加样、孵育、洗涤后,加上Cy3荧光标记物,用激光芯片扫描系统对蛋白质微阵列进行扫描成像。将获得的图像用Imagene专用分析软件进行分析,所获数据根据cutoff值自动生成判断结果。用此蛋白微阵列系统检测了100份4个项目皆阴性的血清标本,确定了其cutoff值。检测了经酶联免疫吸附试验(ELISA)筛选出4个项目皆阴性的标本40例;艾滋病病毒抗体阳性标本30例;乙肝表面抗原、丙肝抗体、梅毒特异性抗体阳性血清各100份。〔结果〕蛋白质微阵列法检测艾滋病病毒抗体、丙肝抗体和乙型肝炎表面抗原的阳性和阴性标本的符合率皆为100%,梅毒特异性抗体阳性符合率为97%,阴性符合率为100%。〔结论〕蛋白质微阵列法检测HIV、HCV、TP和HBsAg与ELISA法检测结果具有高度的符合率。该法具有高通量、快速、特异性强、操作简便等特点,适用于大规模样本的分析,在卫生检疫传染病检测方面具有应用和推广价值。

Objective To establish a protein microarray method that can detect HIV virus （HIV） antibody, hepatitis C virus （HCV） antibodies, syphilis （TP） antibodies and hepatitis B virus surface antigen （HBsAg） at the same time in human serum. Method Firstly HIV, HCV,TP recombinant antigen and hepatitis B virus monoclonal antibody of the genetic engineering were printed onto the surface of activated slides, then the protein micro-array were made up. After the serum samples were diluted, sampled, incubated and washed, then marked by the fluorescent marker Cy3, the protein micro-array were scaned and imaged by a high-resolution laser confocal micro-array scanning analysis system. The images were analyzed and assessed by the Imagine soft ware, and the qualitative results of these samples were got according to their value of the ＂cutoff＂. 100 cases of specimens with four test items all negative were detected by this protein micro-array method, and their results of ＂cutoff＂ value were obtained. 40 cases of specimens with four test items all negative, 30 cases of specimens with anti-HIV positive, every 100 cases of specimens with anti-HCV, anti-TP and HBsAg positive were detected. Result Detecting HIV, HCV, TP antibody and HBsAg by protein micro-array and comparing with ELISA, the coincidences of all negative samples was 100%, the coincidences of all anti-HCV, anti-HIV and HBsAg positive samples was 100%, the coincidences of anti-TP positive samples was 97%. Conclusion The protein micro-array method has high-throughput, speediness, specificity, easy to operate etc, which can apply to large-scale sample analysis, deserves extended application in the detection of infectious diseases of hygiene quarantine.