甲型流感H1N1(2009)流感病毒M2e单克隆抗体的制备及双抗体夹心ELISA检测方法的建立

Preparation of monoclonal antibodies and establishment and development of methods to detect M2E of influenza A H1N1 influenza virus(2009)

目的制备甲型流感H1N1(2009)流感病毒M2e单克隆抗体,并建立检测M2e的双抗体夹心ELISA方法。方法采用碳化二亚胺法将人工合成的M2e多肽分别与牛血清白蛋白(BSA)和卵清蛋白(OVA)进行偶联后作为免疫原,利用杂交瘤细胞技术制备抗A型流感M2e单克隆抗体并进行抗体的鉴定,建立检测M2E的双抗体夹心ELISA方法。结果制备了3株甲型流感H1N1(2009)流感病毒M2e单克隆抗体,经鉴定单克隆抗体均为IGg1a亚型。利用制备的M2e单克隆抗体建立和优化甲型流感M2e检测双抗体夹心ELISA方法,特异性符合临床要求,灵敏度为19ng/ml。结论制备了甲型流感H1N1(2009)M2e单克隆抗体,建立的M2e检测双抗体夹心ELISA方法具有较高的灵敏度和特异性,为研究M2e单克隆抗体作用机制奠定了基础。

Objectives To develop monoclonal antibodies against influenza virus M2e and to establish a method of detec- ting M2e using a double antibody sandwich enzyme-linked immunosorbent assay. Methods A carbodiimide reaction was used to conjugate synthetic M2e peptide with BSA and OVA as an immunogen. The hybridoma technique was used to pre pare and identify monoclonal antibodies to influenza A M2e. Results Three types of monoclonal antibodies to M2e were prepared, and these antibodies were identified as the IGgla subtype. A method of detecting M2e of the influenza A H1N1 virus （2009） was established. This method had a sensitivity of 19 ng/ml. Conclusion This study has laid the founda- tion for the development of kits to detect a range of influenza A antigens, and the current findings can be used to identify influenza caused by the influenza A virus in the future in order to guide clinical treatment.