摘要
本文介绍一种端粒酶检测方法，该方法是将 Ｔ Ｒ Ａ Ｐ法在 Ｐ Ｃ Ｒ 过程中引入３ Ｈｄ Ｃ Ｔ Ｐ，结合液闪计数ｃｐｍ 半定量分析端粒酶活性。应用该方法检测端粒酶阳性的 Ｃ Ｎ Ｅ２ 细胞和部分组织标本及 Ｒ Ｎａｓｅ Ａ 或加热预处理后的对照标本，并与 Ｔ Ｒ Ａ Ｐ 法相比较，结果显示 Ｃ Ｎ Ｅ２ 细胞端粒酶阳性，１０ 个 Ｃ Ｎ Ｅ２ 细胞中仍可检到端粒酶，组织样品的端粒酶与文献值基本一致；放射性活性计数（ｃｐｍ） 与 Ｃ Ｎ Ｅ２ 细胞抽提液中端粒酶活性具有良好的线性关系；用 Ｒ Ｎａｓｅ Ａ 或加热处理后标本为阴性，ｃｐｍ 值接近阴性对照；批内和批间变异度分别为１１ ．６８ ％ 和２０９９ ％ 。该方法不需使用聚丙烯酰胺凝胶电泳（ Ｐ Ａ Ｇ Ｅ） 和放射自显影，简便快速，当天可观察结果，并具有灵敏度高、特异性和批内重复性好之特点。
This study developed a semi quantitative method of telomerase activity. The method was based on the combination of TRAP assay and scintillation count assay(\+3 H dCTP incorporation). It was used to quantify telomerase activity in CNE\-2 cells and some tissue specimens. RNase pretreated or heat treated cells were used as controls. The results demonstrated that telomerase activity measured by this method was positive in CNE\-2 cells, and it could be clearly detected with as few as 10 cells. There was a linear correlation between the radioactive count and the telomerase activity. The telomerase activity of some tissue specimens were accordant with reported data. Telomerase activity of RNase pretreated or heat treated cells was negative, their radioactive counts were almost the same as lysis buffer control. The variations within group and between groups were 11.68% and 20.99%, respectively. This method was free of PAGE and radioautography, and hence simple and fast. The results could be obtained with one day. It showed high sensitivity, good specificity and repeatability.
出处
《癌变·畸变·突变》
CAS
CSCD
1999年第5期213-216,共4页
Carcinogenesis,Teratogenesis & Mutagenesis
基金
美国中华医学会( C M B) 及国家自然科学基金
关键词
端粒酶
定量分析
TRAP法
液体闪烁计数
肿瘤
Telomerase
Quantitative Analysis
Telomeric Repeat Amplification Protocol
Scintillation Count Assay
