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目的探讨不同浓度的PHA、ConA及CD3ε单克隆抗体对Balb/c小鼠淋巴细胞迁移及分泌趋化因子的影响,初步了解这3种刺激剂与小鼠淋巴细胞迁移功能之间的关系。方法通过研磨法分离Balb/c小鼠脾脏淋巴细胞,用不同浓度的PHA、ConA及CD3ε单克隆抗体分别刺激培养淋巴细胞,收集细胞上清,用Traswell法测定细胞上清对淋巴细胞迁移率的影响;ELISA法检测细胞上清中趋化因子XCL1、CCL4、CCL5、CCL3、IL-16、IL-8及CCL20的水平。结果 PHA组各个浓度刺激下淋巴细胞的迁移率和对照组无明显差异;ConA组的刺激浓度为20μg/ml时,其对应的淋巴细胞迁移率明显高于对照组;CD3ε单抗组,其浓度分别为2.5、10μg/ml时,对应的淋巴细胞迁移率明显高于对照组。在各个刺激组中,ELISA测得的多种趋化因子水平在总体上存在明显差异。结论 ConA及CD3ε单克隆抗体分别刺激小鼠淋巴细胞后,其分泌趋化因子的功能发生了明显的改变,这些趋化因子影响着小鼠淋巴细胞的迁移功能。趋化因子XCL1、CCL4、CCL5、CCL3的水平明显高于对照,说明这几个因子在淋巴细胞迁移中起到重要的作用。根据实验结果初步推荐CD3ε单抗可以作为小鼠淋巴细胞迁移试验中的首选阳性对照,次选为ConA。
Objective To investigate the effects of different concentrations of monoclonal antibodies against PHA, ConA and CD3ε on the migration and secretion of chemokines in Balb / c mice lymphocytes and to investigate the relationship between the three stimulators and the migration of mouse lymphocytes. Methods The spleen lymphocytes of Balb / c mice were isolated by grinding method. The lymphocytes were stimulated with different concentrations of PHA, ConA and CD3ε monoclonal antibodies respectively. The supernatants of the cells were collected. The cell viability was measured by Traswell method. The levels of chemokines XCL1, CCL4, CCL5, CCL3, IL-16, IL-8 and CCL20 in the cell supernatant were detected by ELISA. Results There was no significant difference in lymphocyte migration between PHA group and the control group. When the concentration of ConA group was 20μg / ml, the corresponding lymphocyte migration rate was significantly higher than that of the control group. CD3ε mAb group, Respectively, 2.5, 10μg / ml, the corresponding lymphocyte migration was significantly higher than the control group. There was a clear overall difference in the levels of various chemokines measured by ELISA in each stimulation group. Conclusion ConA and CD3ε monoclonal antibodies stimulate the lymphocytes of mice, respectively, after the function of its secretion of chemokines have changed significantly, these chemokines affect the mouse lymphocyte migration. The levels of chemokines XCL1, CCL4, CCL5 and CCL3 were significantly higher than that of the control, indicating that these factors play an important role in lymphocyte migration. Based on the results of the experiment, CD3ε monoclonal antibody was initially recommended as the positive control for mouse lymphocyte migration test, and the second was ConA.