摘要
为了研制鼠抗人BTLA功能性单克隆抗体,以高表达人BTLA分子的基因转染细胞L929/BTLA为免疫原,常规免疫BALB/c小鼠,采用B淋巴细胞杂交瘤技术进行细胞融合,并以基因转染细胞293T/BTLA和293T/mock作为抗原,筛选阳性杂交瘤克隆,经间接免疫荧光标记和流式细胞术分析、反复鉴定和多次克隆化培养,筛选获得分泌特异性鼠抗人BTLA分子单克隆抗体的杂交瘤细胞株;采用Ig亚型快速定性试纸法、细胞核染色体计数、竞争结合抑制试验和T增殖抑制试验等对单抗进行生物学特性的鉴定。结果表明,成功获得一株持续、稳定分泌鼠抗人BTLA单克隆抗体的杂交瘤细胞株8H9,该单抗可特异性识别基因转染细胞以及静止与活化T淋巴细胞上表达的BTLA分子,单抗8H9和BTLA交联后能显著抑制鼠抗人CD3单抗对T细胞激发的增殖作用。
To prepare a functional monoclonal antibody of mouse anti-human B and T lymphocyte attenuator （BTLA）, highly expressed human BTLA transfectant L929/BTLA was used as immunogen to immunize 6-8 weeks BALB/c mice and then the mouse splenic cells were fused with mouse plasmocytoma cells SP2/0. The hybridoma cells were screened with another human BTLA transfectant 293T/BTLA. First strip analysis was performed to identify immunoglobulin subclass of the produced mono- clonal antibodies, and the inhibitory effect on T cell proliferation induced by anti-CD3 mAb of such monoclonal antibodies was studied by MTT assay. The results showed that this newly generated mAb 8H9 could specifically recognize the BTLA protein expressed on these transfected cell lines, and could bind with not only resting but also activated T lymphocytes. Moreover, this monoclonal antibody could mediate obviously an inhibitory effect on the proliferation of T cells.
出处
《现代免疫学》
CAS
CSCD
北大核心
2007年第1期18-22,共5页
Current Immunology
基金
国家自然科学基金重点资助项目(30330540)
国家自然科学基金面上资助项目(30600548)
江苏省"135"临床免疫学重点实验室资助项目
江苏省高校自然科学基础研究资助项目(06KJB310093)
苏州大学医学发展基金资助项目(2006)
