论文部分内容阅读
目的体外表达ＣａｇＡ蛋白，发展快速而简便的检测ｃａｇＡ＋Ｈｐ感染的免疫学方法．方法用ＰＣＲ技术克隆ｃａｇＡ基因５端片段（８５４ｂｐ）于ｐＢＶ２２０中，在ＤＨ５α中温度诱导表达Ｍｒ３８０００的单体ＦＣａｇＡ，复性后分别作阴离子交换及凝胶柱层析，Ｗｅｓｔｅｒｎｂｌｏｔ鉴定ＦＣａｇＡ的抗原性．以重组ＦＣａｇＡ为抗原，通过制备胶体金及免疫胶体金，建立了斑点金免疫渗滤试验（ＤＩＧＦＡ）快速检测ｃａｇＡ＋Ｈｐ感染的方法．结果ＦＣａｇＡ具有抗原性．以该抗原所建立的斑点金免疫渗滤试验（ＤＩＧＦＡ）快速检测ｃａｇＡ＋Ｈｐ感染的方法仅需２ｍｉｎ～３ｍｉｎ可完成，无需特殊设备，并可单人份操作．经过与ＥＩＡ法对比检测２６２例患者血清，本方法的特异性为９８５％，敏感度为９６８％．结论具有抗原活性的ＦＣａｇＡ蛋白的表达及ＤＩＧＦＡ检测抗ＣａｇＡ抗体方法的建立，对ＣａｇＡ＋Ｈｐ感染的流行病学研究具有重要意义
Objective To express CagA protein in vitro and to develop a rapid and simple immunological method for the detection of cagA + Hp infection. Methods The 5? -terminal fragment (854bp) of cagA gene was cloned into pBV220 by PCR. The recombinant plasmid was induced to express Mr38000 in DH5α. After renaturation, the recombinant plasmids were separated by anion exchange and gel filtration chromatography. Western blot was used to identify FCagA antigen Sex. Using recombinant FCagA as antigen, a method of rapid detection of cagA + Hp infection by dot immunogold filtration assay (DIGFA) was established by preparing colloidal gold and immunogold gold. Results FCagA was antigenic. The antigen-based Blot Immunofiltration (DIGFA) assay for rapid detection of cagA + Hp infections takes only 2 to 3 minutes, eliminating the need for special equipment and operating in single doses. After comparing with the EIA method to detect the serum of 262 patients, the specificity of this method was 98.5% and the sensitivity was 96.8%. Conclusion The expression of FCagA with antigenic activity and the detection of anti-CagA antibody by DIGFA are important for the epidemiological study of CagA + Hp infection