活化相关分泌蛋白1单克隆抗体的制备及其在保守结构域鉴定中的应用

目的:制备重组活化相关分泌蛋白1(ASP-1)的单克隆抗体,并用其鉴定保守结构域。方法:用原核表达并纯化的重组ASP-1不加佐剂免疫BALB/c小鼠,采用杂交瘤技术及有限稀释传代法筛选稳定分泌特异性抗体的杂交瘤细胞株,制备单抗腹水后用间接ELISA进行抗体特异性鉴定和效价检测,利用肽结合ELISA和Western印迹鉴定单抗识别的保守结构域。结果:获得5株能稳定分泌抗ASP-1单克隆抗体的杂交瘤细胞株,且5株单抗的识别区域均为21~28氨基酸残基的保守性结构域。结论:制备了抗ASP-1的单克隆抗体,为深入研究ASP-1佐剂的活性功能区及作用机制提供了有效工具。

Preparation and the Conserved Domains Recognition of Monoclonal Antibodies to the Activation-Associated Secreted Protein-1

Objective： To prepare monoclonal antibodies（mAb） specific to recombinant activation-associated secret ed protein-l（ASP-1） and identify the conserved domains. Methods： .The BALB/c mice were immunized with re combinant ASP-1 from prokaryotic expression system without any adjuvant, and the mAb to ASP-1 were generat ed by conventional methods. Antibody titers and specificities of mAb were measured by indirect ELISA. Further more, the conserved domains of ASP-1 were identified by peptide-based ELISA and Western blot. Results： Five strains of hybridoma eel1 lines with the stable secretion of ASP-1 specific mAb were obtained, and these 5 stains of mAb recognized the conserved domain of 21 to 28 amino acid residues. Conclusion： The specific mAb were successfully prepared, and supply an effective tool to study the adjuvanticity domain and mechanism of ASP-1.