原位细胞凋亡的荧光、酶免疫化学法检测实验研究

A Study of In Situ Detection of Cell Apoptosis by Fluorescence and Colorimetric TUNEL

用荧光法和酶免疫化学法末端转移酶介导的ｄＵＴＰ缺口末端标记（ＴＵＮＥＬ）实验分别对小鼠心肌组织、人慢性肝炎肝组织和内膜增生的小型猪血管组织进行了细胞凋亡检测。结果显示：①心肌无凋亡细胞发现，肝组织和血管内膜组织有散在的０～１２个细胞／２５倍物镜视野的细胞标记阳性（凋亡）。②酶免疫化学法ＴＵＮＥＬ实验的检测敏感度较荧光法ＴＵＮＥＬ实验的敏感度高。③过高的末端转移酶可导致假阳性的产生，对荧光法ＴＵＮＥＬ实验，末端转移酶／标记缓冲液比例应为１∶９～１９。

Apoptosis of rat heart muscle, liver tissue in human chronic hepatitis and proliferative plaque of minipig′s iliac arteriae were tested by methods of fluorescence TUNEL and colorimetric TUNEL (terminal deoxynucleotidyl transferasemediated dUTP nick endlabeling). The results showed: ①No apoptosis was found in normal rat myocardium, scattered apoptotic nuclei were labeled in the tissues of liver and ateriae (0￣12 cells per 25 folds objective lens field). ②The sensitivity of colorimetric TUNEL was higher than that of the fluorescence TUNEL. ③Mislabeling (falsepositive) may occur, if the title of terminal deoxynucleotidyl transferase (TdT) in labeling mix was too high, the best ratio of TdT/labeling buffer for fluorescence TUNEL was 1∶9￣19, and for colorimetric TUNEL 1∶59 according to our experiment findings.