de Carvalho H F, Taboga S R
Department of Cell Biology, UNICAMP, Campinas SP, Brazil.
Histochem Cell Biol. 1996 Dec;106(6):587-92. doi: 10.1007/BF02473274.
We have studied the possibility of associating fluorescence microscopy and hematoxylin-eosin staining for the identification of elastic fibers in elastin-rich tissues. Elastic fibers and elastic laminae were consistently identified by the proposed procedure, which revealed itself to be easy and useful for the determination of such structures and their distribution. The fluorescence properties of stained elastic fibers are due to eosin staining as revealed by fluorescence analysis of the dye in solution, with no or only minor contribution by the elastin auto-fluorescence. The main advantage of this technique resides in the possibility of studying the distribution of elastic fibers in file material without further sectioning and staining. The use of the confocal laser scanning microscope greatly improved the resolution and selectivity of imaging elastic fibers in different tissues. The determination of the three-dimensional distribution and structure of elastic fiber and laminae using the confocal laser scanning microscope was evaluated and also produced excellent results. used to discriminate calcified bone and elastic fibers using the fluorescence microscope (Bradbeer et al. 1994). The selective fluorescence exhibited by these two main tissue components is based on the very faint eosin staining, while intensely eosinophilic substrates and those stained with the highly absorbing basic dyes show no fluorescence or just a very faint signal. Considering the possibility of using eosin fluorescence for the selective identification of some tissue components, we have extended this approach to the study of elastic fibers in hematoxylin-eosin (H&E) preparations. This paper then describes the results of observations with the fluorescence microscope of some H&E stained sections and evaluates the use of the confocal laser scanning microscope to image elastic fibers in the same preparations. Furthermore, we have also studied absorption and fluorescence spectra of eosin and phloxine, a closely related dye, in solution, to correlate them with the fluorescence detected in tissue sections.
我们研究了将荧光显微镜检查与苏木精 - 伊红染色相结合以鉴定富含弹性蛋白组织中弹性纤维的可能性。通过所提出的程序能够始终如一地识别弹性纤维和弹性板层，该程序对于确定此类结构及其分布而言简便且实用。染色弹性纤维的荧光特性归因于伊红染色，这通过对溶液中染料的荧光分析得以揭示，弹性蛋白自身荧光的贡献极小或几乎没有。该技术的主要优势在于能够在无需进一步切片和染色的情况下研究存档材料中弹性纤维的分布。共聚焦激光扫描显微镜的使用极大地提高了对不同组织中弹性纤维成像的分辨率和选择性。利用共聚焦激光扫描显微镜对弹性纤维和弹性板层的三维分布及结构进行了测定，结果也非常出色。曾有人使用荧光显微镜鉴别钙化骨和弹性纤维（布拉德比尔等人，1994年）。这两种主要组织成分所呈现的选择性荧光基于非常微弱的伊红染色，而强嗜伊红性底物以及用高吸收性碱性染料染色的底物则无荧光或仅有非常微弱的信号。考虑到利用伊红荧光选择性鉴定某些组织成分的可能性，我们将此方法扩展至对苏木精 - 伊红（H&E）染色制剂中弹性纤维的研究。本文随后描述了用荧光显微镜观察一些H&E染色切片的结果，并评估了在相同制剂中使用共聚焦激光扫描显微镜对弹性纤维成像的情况。此外，我们还研究了溶液中伊红和与之密切相关的染料焰红的吸收光谱和荧光光谱，以将它们与在组织切片中检测到的荧光相关联。
