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目的:建立可特异识别HL60和HL60/ADR细胞膜表面差异蛋白的抗体库,制备单克隆抗体并研究其生物学活性。方法:通过Cp诱导的消减免疫法免疫BALB/c小鼠,采用传统杂交瘤技术制备可特异识别两种细胞差异膜蛋白的单克隆抗体;采用FACS和激光共聚焦显微镜鉴定其和两种靶细胞结合的特异性和差异性。结果:获得51株候选的差异抗体,成功筛选并纯化其中一株单抗(5F6)。5F6与可特异稳定的识别HL60/ADR细胞,结合率扣除本底为68.2%,而该抗体与HL60细胞的结合率为17%。结论:SI结合差异筛选的方法是制备差异抗体便捷可行的方法,所获得的单克隆抗体与HL60和HL60/ADR细胞膜蛋白结合有特异性和差异性,实验中所获得的单抗是寻找肿瘤耐药的新靶点和发现肿瘤耐药新机制的工具,为研究这些问题开拓了新思路。
OBJECTIVE: To establish an antibody library that can identify differentially expressed proteins on the surface of HL60 and HL60 / ADR cells and prepare monoclonal antibodies to study their biological activity. Methods: BALB / c mice were immunized with Cp-induced immunosuppression. Monoclonal antibodies that could specifically recognize two different cell membrane proteins were prepared by using the hybridoma technique. FACS and confocal laser scanning microscopy were used to identify the two monoclonal antibodies against the two target cells Specificity and diversity of binding. Results: Fifty-one candidate differential antibodies were obtained and one of the McAbs (5F6) was successfully screened and purified. 5F6 and specifically stably recognize HL60 / ADR cells, the binding rate deducted background was 68.2%, and the binding rate of the antibody to HL60 cells was 17%. CONCLUSIONS: The method of SI binding and differential screening is a convenient and feasible method for preparing differential antibodies. The obtained monoclonal antibodies have specificity and difference in binding to the cell membrane proteins of HL60 and HL60 / ADR. The monoclonal antibodies obtained in the experiment are tumor-resistant New targets of medicine and tools to discover new mechanisms of drug resistance in cancer have opened up new ideas for the study of these problems.