间接ELISA法测定转基因小鼠血清中核糖核酸酶抑制因子的表达

Expression of ribonuclease inhibitor in blood serum of RI gene infected mice by indirect ELISA technique

关键词:  核糖核酸酶抑制因子  间接酶联免疫吸附试验  血管生长因子

Key words:  ribonuclease inhibitor (RI)  indirect enzyme linked immune-osorbent assay (ELISA)  angiogenin（Ang）

摘要:

［目的］建立间接ELISA法测定血清中核糖核酸酶抑制因子（ribonuclease inhibitor, RI）含量的方法，并测定在转RI基因小鼠血清中RI的表达情况。［方法］ 用血清样品包被酶标板，兔抗人 RI抗体为一抗，辣根过氧化物酶标记的羊抗兔IgG为二抗，建立血清RI定量检测的间接ELISA法；取转染了RI基因的小鼠的血清，用上述建立的间接ELISA法测定不同时相小鼠血清中RI的表达水平。［结果］ 定量检测血清RI的间接ELISA法最适反应条件为一抗最佳稀释度为1∶4000，二抗的最佳稀释浓度为1∶2000。转基因小鼠第2、3周血清中RI含量较低（0.556±0.080），1个月左右达到最高值（0.836±0.113），3个月时明显下降（0.640±0.100）。［结论］ 间接ELISA法可以用于测定血清中的RI抗原含量。

Abstract:

［Objective］To establish indirect ELISA method for detection of ribonuclease inhibitor (RI) antigen in serum and to analyze the dynamic change of RI in the blood of mice transinfected with RI gene. ［Method］ The RI in serum was used as antigen, rabbit anti-RI was used as the first antibody, and goat anti-rabbit IgG-HRP was used as the second antibody. The best dilution for rabbit anti-RI and goat anti-rabbit IgG-HRP were decided according to the optimal reaction condition standards for ELISA, the indirect ELISA for detection of RI antigen expressed in serum was established. By this method, the RI antigen in serum of mice which were transinfected with RI gene was tested in different periods. ［Result ］ The best dilution of first and second antibody were 1∶4 000 and 1∶2 000 respectively. The RI antigen in mice serum reached peak at one month, then decreased. ［Conclusion］ The indirect ELISA was a specific and sensitive method in testing the expression of RI antigen and could be used to analyze the dynamic change of RI in serum.