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为检测凋亡过程中活细胞染色质致密程度而建立荧光测量系统 :方法 :2 4孔培养板内置盖玻片 ,培养食管癌细胞 ,加入 3μmolAs2 O3诱导凋亡 ,6 ,12 ,2 4小时后 ,实验组和对照组用活体染料Hoechst 33342染色 ,落射荧光显微镜配相差装置观察。显微镜下装有冷CCD检测系统 ,测定荧光强度(FI) ,用计算机StarⅠ软件分析。同时在同样处理条件下 ,另一标本上作原位末端标记 (TUNEL)荧光检查。结果 : 加入As2 O3诱发细胞凋亡 ,细胞形态改变为卵圆形 ,细胞核皱缩、固缩或核碎裂凋亡的改变。FI显示细胞核染色质致密程度逐步增加。同时TUNEL反应阳性 ,证明有DNA链断裂。结论 : 此方法可以检测活细胞凋亡时染色质致密程度的动态变化 ,可望作为诊断凋亡细胞的指标 ,也可用于区别常染色质和异染色质。
A fluorescence measuring system was set up to detect the chromatin density of living cells during apoptosis: Methods: A 2 4-well culture plate was covered with glass coverslips to induce esophageal cancer cells. 3μmol of As 2 O 3 was added to induce apoptosis. After 6, 12, 24 hours , Experimental group and control group with live dye Hoechst 33342 staining, epifluorescence microscope with phase difference device observation. The microscope was equipped with a cold CCD detection system to measure fluorescence intensity (FI) and analyzed using computer StarⅠ software. At the same time under the same treatment conditions, another specimen for end-TUNEL fluoroscopy. Results: Apoptosis was induced by adding As2 O3, and the morphological changes were oval, nuclear shrinkage, pyknosis or nuclear fragmentation. FI showed a gradual increase in chromatin density in the nucleus. At the same time TUNEL reaction was positive, showing that DNA strand breaks. Conclusion: This method can detect the dynamic changes of chromatin density when living cells are apoptosis, which is expected to be used as an index to diagnose apoptotic cells and to distinguish between eosinophilic and heterochromatic.