Curtis S K, Cowden R R
Histochemistry. 1981;72(1):39-48. doi: 10.1007/BF00496777.
Microfluorometric estimates of total RNA were made in selected test material stained with berberine sulfate, acridine orange, and Hoechst 33258. These measurements were compared with those obtained with propidium iodide, which is known to interact only with double-stranded nucleic acids. It was observed that all of the fluorochromes, including propidium iodide, yielded very similar patterns of fluorescence in the various types of material tested. In isolated thymocyte and hepatocyte nuclei stained with either propidium iodide or Hoechst 33258 at pH 2, it was evident that RNA could be estimated only indirectly by measuring the amount of fluorescence before and after extraction with RNase. It was apparent that the total fluorescence of small thymocyte nuclei was affected much less by RNase extraction than that of 2c hepatocyte nuclei. Attempts to obtain direct estimates of RNA by exposing the preparations to DNase were not successful: the fluorescence of thymocyte nuclei dropped almost to zero, and hepatocyte nuclei could no longer be assigned to distinct ploidy classes. In addition, since the highly condensed chromatin of thymocyte nuclei was stained much more prominently than the looser chromatin of hepatocyte nuclei with Hoechst 33258, it was apparent that this fluorochrome - when used at pH 2 - has potential usefulness as a "probe" of organizational differences in chromatin.
采用微荧光法对用硫酸黄连素、吖啶橙和Hoechst 33258染色的选定测试材料中的总RNA进行了估算。将这些测量结果与用碘化丙啶获得的测量结果进行了比较，已知碘化丙啶仅与双链核酸相互作用。观察到，包括碘化丙啶在内的所有荧光染料在各种测试材料中产生的荧光模式非常相似。在用碘化丙啶或Hoechst 33258在pH 2条件下染色的分离胸腺细胞核和肝细胞核中，很明显，只有通过测量用核糖核酸酶提取前后的荧光量才能间接估算RNA。很明显，小胸腺细胞核的总荧光受核糖核酸酶提取的影响比2c肝细胞核小得多。通过将制剂暴露于脱氧核糖核酸酶来直接估算RNA的尝试未成功：胸腺细胞核的荧光几乎降至零，肝细胞核不再能归为不同的倍体类别。此外，由于用Hoechst 33258染色时，胸腺细胞核高度浓缩的染色质比肝细胞核松散的染色质染色更明显，很明显，这种荧光染料在pH 2条件下使用时，有可能作为染色质组织差异的“探针”。
