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〔目的〕针对甲型流感病毒血凝素(Hemagglutinin,HA)蛋白同一区段不同亚型的合成肽,比较分析其免疫反应性和交叉反应性,获取具有免疫诊断学意义的抗原肽。〔方法〕通过生物信息学分析,针对新甲型H1N1、季节性H1N1、H2N2、H3N2和H5N1共5种亚型甲型流感病毒,选择2个HA肽段区位设计2组抗原肽段(19肽和23肽)合成,每组5条共10条,并分别与钥孔戚血蓝蛋白(KLH)和牛血清白蛋白(BSA)载体偶联形成全抗原后,免疫家兔制备兔源抗体,建立免疫反应性筛选的酶联免疫吸附法(ELISA),比较合成肽的免疫反应性。〔结果〕以阳性血清平均OD490nm/阴性血清平均OD490nm(P/N)值为判断标准,筛选出具有免疫反应性的2条特异性抗原肽(KYVKSTKLRLAT-GLRNVPS,VKNGTYDYPQYSEEARLNREEIS),前者具有亚型特异反应性,后者则具有一定的亚型通用免疫反应性。〔结论〕筛选出具有免疫反应性合成肽抗原,为进一步建立流感病毒亚型的快速鉴定诊断方法奠定了基础。
[Objective] To study the immunological reactivity and cross-reactivity of synthetic peptides with different subtypes of the same segment of the influenza A virus Hemagglutinin (HA) protein, and to obtain antigenic peptides of immunological diagnostic significance. [Methods] According to bioinformatics analysis, two groups of antigenic peptides (19 peptides were designed according to the five HA subtype A (H1N1), seasonal H1N1, H2N2, H3N2 and H5N1 influenza A viruses And 23 peptide) were synthesized. Each group consisted of 10 plasmids in total of 10 and were conjugated with keyhole limpet hemocyanin (KLH) and bovine serum albumin (BSA) to form rabbit anti-rabbit antibodies Immunoreactive screening of enzyme-linked immunosorbent assay (ELISA) to compare the immunological reactivity of the synthesized peptides. [Results] Two specific antigenic peptides (KYVKSTKLRLAT-GLRNVPS, VKNGTYDYPQYSEEARLNREEIS) with immunoreactivity were screened based on the average serum OD490nm / OD490nm (P / N) value of the serum and the former had subtype-specific responses While the latter has a certain subtype of universal immunoreactivity. 〔Conclusion〕 Screening out immunoreactive synthetic peptide antigen, which laid the foundation for the rapid identification and diagnosis of influenza virus subtypes.