Santi P, Papa S, del Coco R, Falcieri E, Zini N, Marinelli F, Maraldi N M
Biol Cell. 1987;59(1):43-54. doi: 10.1111/j.1768-322x.1987.tb00514.x.
Ethidium bromide (EB) is widely used for investigating the DNA conformation in chromatin both with conventional and cytofluorimetric techniques. Since the interaction of the dye with DNA should result in structural deformations which can be different in isolated or in situ chromatin, a study has been performed on the effects caused by different amounts of EB and the analogous propidium iodide on isolated nuclei, in which chromatin maintains its native relationships with the other nuclear structures (envelope, nucleolus, interchromatin RNP, nuclear matrix). The results obtained by comparing ultrastructural observations in thin sections and in freeze-fracturing with conformational analysis in multiparameter flow cytometry indicate that the phenanthridinic fluorochromes, especially at the high concentrations used for cytofluorimetric analyses, cause deep rearrangements of the chromatin in situ. These effects consist both in aggregation and condensation of the fibers into the dense chromatin domains, and in an increase of the supernucleosomal configuration associated with an enlargement of interchromatin spaces in which the RNP particles appear particularly evident. These results, discussed with those available on isolated chromatin, suggest that any unwinding effect of the intercalating dyes on the DNA cause a general condensation of chromatin as a consequence of the constraints which characterize the organization of the chromatin inside the nucleus.
溴化乙锭（EB）广泛应用于采用传统技术和细胞荧光测定技术研究染色质中的DNA构象。由于该染料与DNA的相互作用会导致结构变形，而这种变形在分离的染色质或原位染色质中可能有所不同，因此我们进行了一项研究，探究不同量的EB以及类似物碘化丙啶对分离细胞核的影响，在这些细胞核中，染色质与其他核结构（核膜、核仁、染色质间核糖核蛋白、核基质）保持着天然的关系。通过比较超薄切片和冷冻断裂的超微结构观察结果与多参数流式细胞术中的构象分析结果发现，菲啶类荧光染料，尤其是在用于细胞荧光分析的高浓度下，会导致原位染色质发生深度重排。这些影响既包括纤维聚集和凝聚形成致密的染色质结构域，也包括超核小体构型增加，同时伴随着染色质间空间扩大，其中核糖核蛋白颗粒显得尤为明显。将这些结果与关于分离染色质的现有结果进行讨论后表明，嵌入染料对DNA的任何解旋作用都会导致染色质普遍凝聚，这是由于细胞核内染色质组织所具有的限制因素所致。
