Three-dimensional reconstructions from non-deparaffinized tissue sections
Three-dimensional reconstructions from non-deparaffinized tissue sections
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非脱蜡组织切片的三维重建
DOI：
10.1007/s00429-005-0006-8
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发表时间：
2005
期刊：
影响因子：
--
通讯作者：
L. Kubínová
中科院分区：
文献类型：
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作者：
M. Jirkovská;I. Náprstková;J. Janáček;T. Kučera;J. Macášek;P. Karen;L. Kubínová
显微图像的三维重建是胚胎学和发育生物学中研究生物结构的一个有用工具。然而，通常需要科普与样品制备有关的许多困难。为了最小化连续切片中结构的相互位移，测试了非脱蜡组织切片用于3-D重建的适用性。将鸡胚固定并用伊红完全染色，然后包埋在石蜡中。约30 μ m厚的未脱蜡连续切片用于获得初始数据，用于使用荧光或共聚焦显微镜对较大堆叠的胚胎体进行三维重建。同时收集中肾光学连续切片作为其三维重建的源图像，并进行纤维连接蛋白、层粘连蛋白和波形蛋白的免疫组化检测。结果发现，保留石蜡的切片保留了组织成分的相互空间关系，并提供了良好的结构分化。它使三维重建过程更容易。免疫组化反应的产物的本地化表明，共定位的纤维连接蛋白和层粘连蛋白在基底层和波形蛋白的存在下，肾小球和间充质组织。非脱蜡切片的使用代表了用于3-D重建目的的薄组织学切片的耗时更少且更有效的替代方案，并且能够进一步应用材料。
Three-dimensional (3-D) reconstruction from microscopic images represents a useful tool for the study of biological structures in embryology and developmental biology. However, it is usually necessary to cope with many difficulties connected with the preparation of specimens. In order to minimize mutual displacement of structures in successive sections, the applicability of non-deparaffinized tissue sections for 3-D reconstruction was tested. Chicken embryos were fixed and stained in toto with eosin and then embedded in paraffin. About 30-μm-thick non-deparaffinized serial sections were used for obtaining initial data for 3-D reconstruction of larger stacks of embryonic bodies using either fluorescence or confocal microscope. The same sections served for both collecting optical serial sections of mesonephros as source images for its 3-D reconstruction, and immunohistochemical detection of fibronectin, laminin and vimentin. It was found that sections with retained paraffin preserve the mutual spatial relationships of tissue components as well as provide an excellent differentiation of structure. It makes the process of 3-D reconstruction easier. The localization of the products of immunohistochemical reactions demonstrated the co-localization of fibronectin and laminin in basal laminas and the presence of vimentin in glomeruli and mesenchymal tissue. The use of non-deparaffinized sections represents a less time consuming and more effective alternative to thin histological sections for the purpose of 3-D reconstruction, and enables further application of material.