抗人LSECtin单克隆抗体的制备与初步鉴定

Preparation and characterization of monoclonal antibody against human LSECtin

目的：制备抗肝脏、淋巴结窦内皮细胞C型凝集素（LSECtin）单克隆抗体（mAb），并进行特性鉴定。方法：采用原核表达的LSECtin免疫BALB／c小鼠，以间接ELISA法筛选分泌特异性mAb的杂交瘤细胞，采用蛋白印迹、间接免疫荧光、流式细胞术和免疫组化染色法鉴定mAb的特异性。结果：共获得8株可稳定分泌mAb的杂交瘤细胞株。mAb的Ig亚类均为IgG，效价达1：10^6-1：10^7。这些mAb均可识别转染3T3细胞膜上的人LSECtin，6株mAb可特异识别肝脏窦内皮细胞。结论：成功地制备8株抗LSECtin的mAb，经免疫印迹、流式细胞术和免疫组化染色检测，这些mAb的特异性良好，为研究LSECtin的功能提供了有力的试剂。

AIM： To prepare and characterize monoclonal antibody （mAb） against human LSECtin （ liver and lymph node sinusoidal endothelial cell C-type lectin ） protein. METHODS： BALB/c mice were immunized with prokaryotically expressed human LSECtin protein. The splenocytes from the immunized mice were fused with murine myeloma cells （Sp2/0） and then the mAb-positive hybridoma cells were screened by indirect ELISA. Reaction of mAb to LSECtin antigen was characterized by Western blot, indirect immunofluorescent staining, immunohistochemical staining and FCM. RESULTS： Eight hybridoma cells secreting mAbs were established. The isotypes of the mAbs were IgG. Ascites titers were between 1：10^6 -1：10^7. All the mAbs recognized human LSECtin protein on LSECtin-transfected 3T3 cells and six of the mAbs specifically recognized liver sinusoidal endothelial cells. CONCLUSION： Eight anti-LSECtin mAbs have been obtained. The characterization of the mAbs indicate that they show fine specificity by Western blot, indirect immunofluorescent staining, immunohistochemical staining and FCM, which can provide a powerful reagent for the functional study of LSECtin.