Tissue Precise Spatial Structure Determination With Three-Dimensional Reconstruction Technology Based On Ultrathin Sections

Precise structure analysis of biological tissues is the basis ofunderstanding the physiological functions of the organism. The mainmethod is to establish three-dimensional reconstruction of the tissuesamples, including samples processing, sectioning, staining, imaging, andreconstruction. Sections include paraffin sections, frozen sections,semi-thin and ultra-thin slices. Paraffin sections and frozen sections arewidely used in clinical pathological analysis; Semi-thin and ultra-thinsections are generally prepared for the observation of cell microstructurewith electron microscope or AFM, which are thinner and the resolution ismuch higher than that of paraffin and frozen sections. We can get theinterior structure of tissue through reconstruction with the serial sections.Among current methods, there exists a defect that the resolution of Z-axisis much too low.The X, Y-axis direction resolution of current traditional microscope isabout250nm, and the Z-axis direction is about700nm, actually, the realresolution is more than1000nm. Accordingly, the improvement of Z-axisresolution is the key to improve the resolution of microscopic imaging. Inthis paper, we have established a high resolution imaging system with theZ-axis resolution improvement by ultra-thin cutting technology. Tissue was embedded in LR White resin and polymerized, which was cut into50nm-1000nm serial slices with diamond knife. Thereby the Z-axisresolution of50nm-1000nm is obtained. The serial semi-thin or ultra-thinslices are stained with methylation blue and basic fuchsin or labeled withantibody for structure illustration. Three-dimensional reconstruction oftissue can be got using the established high-resolution microscopy imagingtechnology, including2D stitching and3D reconstruction technology,which suppllies much finer structure and have higher resolution thancurrent traditional methods. The established high-resolutionthree-dimensional imaging technique is also applicable to the other tissues,and will provide a solution for ultra-high resolution imaging technology toimprove the Z-axis resolution.