Three-Dimensional Visualization of the Ependyma and Tanycytes in the Brain

The aim of the present work was to develop a complex approach allowing confocal laser microscopy and immunocytochemical reactions for vimentin to be used for spatial reconstruction of the cells lining the cerebral ventricles. Experiments were performed on paraffin sections of the rat brain with different thicknesses (5 and 10 μm). Immunocytochemical reactions were visualized using stains fluorescing in the visible range for selective nuclear staining, i.e., SYTOX Green, and streptavidin-conjugated indocarbocyanin (Cy3). Testing of different processing regimes led to development of a protocol providing intense staining of structures. Use of this set of fluorochromes in confocal laser microscopy allowed easy discrimination between channels and, where required, analysis of structures independently of each other, without the need for an expensive ultraviolet laser. © 2015, Springer Science+Business Media New York.