Ciancio G, Pollack A, Taupier M A, Block N L, Irvin G L
Research Service, Veterans Administration Medical Center, Miami, FL 33125.
J Histochem Cytochem. 1988 Sep;36(9):1147-52. doi: 10.1177/36.9.2457047.
We developed a rapid technique for preservation of Hoechst 33342/propidium iodide-stained cells, using ethanol as a fixative. Combined staining with these dyes makes possible analysis of cell-cycle phase-specific cell death. The technique relies on exclusion of propidium iodide from the viable cells, whereas Hoechst stains all of the cells. The bivariate histograms resulting from the flow cytometric analysis contain the equivalent of two single-parameter DNA histograms, one of the living and the other of the dead cell population. Preservation of staining involved addition of 25% ethanol in PBS after propidium iodide staining and before Hoechst staining. The separation between the living and the dead cell populations was maintained for over 3 days at 4 degrees C. This technique will be valuable for quantitative evaluation of the cell-cycle phase-specific effects of cytostatic or cytotoxic agents, particularly in situations where a lag period between staining and analysis is unavoidable.
我们开发了一种使用乙醇作为固定剂来保存经Hoechst 33342/碘化丙啶染色细胞的快速技术。这两种染料的联合染色使得分析细胞周期阶段特异性细胞死亡成为可能。该技术依赖于活细胞排斥碘化丙啶，而Hoechst可对所有细胞进行染色。流式细胞术分析产生的双变量直方图包含相当于两个单参数DNA直方图，一个是活细胞群体的，另一个是死细胞群体的。染色保存包括在碘化丙啶染色后、Hoechst染色前向PBS中加入25%乙醇。活细胞和死细胞群体之间的分离在4℃下可维持3天以上。该技术对于定量评估细胞周期阶段特异性的细胞生长抑制剂或细胞毒性剂的作用将具有重要价值，特别是在染色和分析之间存在不可避免的延迟期的情况下。
