目的：从人白血病细胞系J6-1克隆并表达变异的M-CSF的功能区，研究其受体结合活性。方法：RT-PCR克隆muM-CSF，并在大肠杆菌BL21trxB(DE3)、pET32c(+)系统中表达；镍柱鏊合层析，抗体亲和层析纯化。ELISA法测定其受体结合活性和解离常数，并测定对J6-1细胞增殖刺激的活性。结果： muM-CSF可在本文实验系统呈可溶性表达，纯化产物具有M-CSF受体的结合活性，解离常数为3.7 nmol/L低于正常M-CSF，且刺激细胞体外增殖能力大于正常M-CSF。结论：从人白血病细胞系J6-1克隆的muM-CSF能在BL21,pET32c(+)系统表达，可得电泳纯产物。

Objective: To clone and express functional part of mutant M-CSF (muM-CSF) from human leukemic cell line J6-1 and investigate its Kd for dissociation and biological activity on the proliferation of J6-1.Method: Functional part of muM-CSF was cloned by RT-PCR and inserted into pET32c(+) and expressed in E.coli BL21trxB (DE3). The recombinant protein was purified through Ni2+ affinity column and antibody linked affinity column. ELISA was performed to define the Kd of the muM-CSF to its receptor. Colony formation assay was performed to test its effects on the proliferation of J6-1. Results: The protein was purified and its Kd to the receptor was 3.7 nmol/L. muM-CSF showed elevated proliferation-stimulating potential than normal M-CSF. Conclusion: muM-CSF could be expressed in the pET32c(+), BL21 system and the muM-CSF showed elevated proliferation promoting ability as to normal M-CSF.

Q786

天津市科技发展计划项目资助（003119311）