鼠抗人FL单克隆抗体的研制及其生物学特性  

Preparation of three functional monoclonal antibodies against human FL molecule and analysis of their bioactivity

作　　者：周春刚[1] 田晓静[1] 居颂光[1] 徐颖[1] 於葛华[1] 张学光[1] 顾宗江[1] 

机构地区：[1]苏州大学医学院免疫学教研室,江苏苏州215007

出　　处：《细胞与分子免疫学杂志》2007年第9期850-852,855,共4页Chinese Journal of Cellular and Molecular Immunology

摘　　要：目的:研制鼠抗人FL单克隆抗体(mAb)以及对其生物学特性的研究。方法:以人FL基因转染细胞株L929-FL为免疫原,采用B细胞杂交瘤技术,获取分泌抗人FLmAb的杂交瘤细胞株。快速定性试纸法鉴定mAb亚类,体内诱生腹水法制备mAb,免疫亲和层析法纯化mAb,MTT法检测mAb对白血病细胞株U937和HL-60生长的影响,Annexin V/PI染色流式细胞术(FCM)检测细胞凋亡。结果:建立了3株稳定分泌抗人FL mAb的杂交瘤,命名为3C2、3C6和8D10。快速定性试纸法鉴定3株mAb均属小鼠lgG2a亚类。3株mAb分别识别白血病细胞U937和HL-60上表达的FL分子。将3株mAb分别加入共表达FL/Flt3的U937和HL-60细胞的培养体系中,MTT法检测显示,3C2、3C6和8D10对白血病细胞的生长均具有抑制作用(P<0.05)。Annexin V/PI染色FCM检测细胞凋亡,3C2和8D10均能增加白血病细胞凋亡的作用。结论:3C2、3C6和8D10是3株功能性抗FL mAb,对于研究FL/Flt3在白血病发生、发展中的作用具有潜在的应用价值。AIM： To Prepare three functional monoclonal antibodies （mAbs） against human FL molecule and analyze their bioactivity. METHODS： The cell line L929-FL transfected with human FL gene was used as immunogen. The hybridomas secreting the antibodies against human FL were obtained by fusing splenoecytes from the immunized mice with murine myeloma cells （Sp2/0）. Their subclasses were analyzed using fast-strip method. The monoclonal antibodies were produced in mouse peritoneal cavity and purified by Protein G affinity chromatography. The inhibitory effect of mAbs against FL on leukemia cell lines U937 and HL-60 was detected by MTT. The apoptosis of U937 and HL-60 cells stained by annexin-V/PI was determined by FCM. RESULTS： Three hybridomas named 3C2, 3C6 and 8D10 were successfully obtained, which secreted monoclonal antibodies against human FL molecule stably. Their subclasses were the mouse IgG2a with kappa light chains. The three monoclonal antibodies recognized the FL molecule on U937 and HL-00 cells that also coexpressed Fit3 molecule. When U937 and HL-60 cells were cultured in presence of 3C2, 3C6 and 8D10, their proliferation was reduced as compared to that in control in MTT assay （P 〈 0.05）. The analysis of annexin-V/PI binding to U937 and HL-60 cells by FCM showed the mAbs had the apoptotogenic activity of the monoclonal antibodies against human FL molecule. CONCLUSION： 3C2, 3C6 and 8D10 are three funtional monoclonal antibodies against human FL molecule. They may be of some value in the study of the roles of FL/FIt3 interaction in leukemia pathogenesis.

关 键 词：FL 单克隆抗体 白血病 

分 类 号：R392.11[医药卫生—免疫学]