ICA512的表达及人抗ICA512抗体测定方法的建立

The cloning and protein expression of ICA512 gene and its clinical application

目的:从人胰脏β-细胞中对ICA512进行基因克隆和蛋白质表达,最终应用于人自身抗体的测定。方法:应用逆转录技术从人胰脏RNA中合成出ICA512的cDNA。经DNA PCR扩增后,克隆到两种带有不同融合蛋白的表达质粒,然后转化至大肠杆菌中,表达出两种该重组蛋白。并利用该两种表达蛋白建立起抗ICA512抗体的ELISA测定法(双抗原夹心法)。结果:用英国RSR ICA512测定试剂盒对表达蛋白进行免疫学分析对照,表明研究中表达的蛋白是具有免疫活性的ICA512。建立的ELISA和RSR放免法的测定结果基本吻合。结论:研究中所表达的ICA512可用于人自身抗体测定进行1型糖尿病(T1DM)诊断,在操作的简便性方面更是优于同类进口测定试剂盒。

Objective：To clone the gene of ICA512 and express its protein from human pancreatic tissue β cell in order to apply for autoimmune antibody assay. Methods：The cDNA of ICA512 was generated by RT-PCR from the RNA extracted from the human pancreatic tissue β cell. After having amplified the DNA by PCR and cloned into replicate plasmid , the replicate plasmid was further cloned into expressing plasmid followed by transfected into the E. coli, two recombinant proteins were expressed. The anti- ICA512 antibody assay method was established by using these expression proteins. Results ： The immune reactivity of the expression protein was conformed by using the ICA512 assay kit （RSR Hd UK）. Conclusion：The expression protein that delineated in the paper can be applied for human anti- ICA512 antibody assay in the diagnosis of T1DM. Technically,the assay system is more easier than the same sort of assay kit on the markets.