CAJ | 학술논문

目的制备鼠抗人IgE高亲和力受体FcεRIα的单克隆抗体并对其进行初步鉴定,为过敏性疾病的相关研究提供条件.方法用人FcεRIα重组蛋白作为抗原免疫BALB/C小鼠,常规方法进行融合,经筛选及克隆化建立可稳定分泌抗人FcεRIα mAb的杂交瘤细胞株,用ELISA、免疫荧光标记、荧光显微镜和流式细胞仪分析以及Western-Blot方法进行抗体的初步鉴定.结果筛选到2株可稳定分泌抗人FcεRIα单克隆抗体的细胞株,流式细胞术分析此2株抗体与CHO3D10细胞的结合率均大于95%,免疫荧光显示2株抗体均能与FcεRIα发生特异性反应.Western-Blot结果显示抗体轻重链分别约28 Kd与50 Kd.结论制备的两株FcεRIα的单克隆抗体可特异性与细胞表面FcεRI结合,可用于肥大细胞功能以及变态反应病临床治疗的进一步研究.
목적 제비서항인IgE고친화력수체FcεRIα적단극륭항체병대기진행초보감정,위과민성질병적상관연구제공조건.방법 용인FcεRIα중조단백작위항원면역BALB/C소서,상규방법진행융합,경사선급극륭화건립가은정분비항인FcεRIα mAb적잡교류세포주,용ELISA、면역형광표기、형광현미경화류식세포의분석이급Western-Blot방법진행항체적초보감정.결과 사선도2주가은정분비항인FcεRIα단극륭항체적세포주,류식세포술분석차2주항체여CHO3D10세포적결합솔균대우95%,면역형광현시2주항체균능여FcεRIα발생특이성반응.Western-Blot결과현시항체경중련분별약28 Kd여50 Kd.결론 제비적량주FcεRIα적단극륭항체가특이성여세포표면FcεRI결합,가용우비대세포공능이급변태반응병림상치료적진일보연구.
Objective To prepare and initially identify the monoclonal antibody against FcεRIα,so as to provide basis for further reseach on anaphylactic diseases.Methods BALB/C mice was immunized with the recombination protein,and the hybridoma cell line which stably secreted FcεRIα mAb was prepared via fusion,screening and cloning.Then ElISA,immunofluorescence labeling method,fluorescent microscopy examination,flow cytometry and Western-Blot assay were adopted to perform initial identification of the monoclonal antibody.Results Two cell strains were screened which stably secreted FcεRIα mAb against FcεRIα. Flow cytometry showed that the binding rate of the antibodies to CH03D10 cells was over 95%.The antibodies named as B4C2 and B584 displayed strong specifility and high affinity.The titration of antibodies in supernatant and mouse aseites were higher than 105.The immunofluorescence assay demonstrated that the antibodies specifically binded with CH03D10 cells,and Western Blot showed the light and heavy chains of the antibodies were 28 Kd and 50 Kd respectively.Conclusion The prepared two strains of specific mAbs against human FcεRIαcan bind with FcεRI on cell surface,which can be used as a tool of further research on mast cell function and clinical therapy of allergosis.