Anaglyph of Retinal Stem Cells and Developing Axons: Selective Volume Enhancement in Microscopy Images

Abstract

Retinal stem cell culture has become a powerful research tool, but it requires reliable methods to obtain high-quality images of living and fixed cells. This study describes a procedure for using phase contrast microscopy to obtain three-dimensional (3-D) images for the study of living cells by photographing a living cell in a culture dish from bottom to top, as well as a procedure to increase the quality of scanning electron micrographs and laser confocal images. The procedure may also be used to photograph clusters of neural stem cells, and retinal explants with vigorous axonal growth. In the case of scanning electron microscopy and laser confocal images, a Gaussian procedure is applied to the original images. The methodology allows for the creation of anaglyphs and video reconstructions, and provides high-quality images for characterizing living cells or tissues, fixed cells or tissues, or organs observed with scanning electron and laser confocal microscopy. Its greatest advantage is that it is easy to obtain good results without expensive equipment. The procedure is fast, precise, simple, and offers a strategic tool for obtaining 3-D reconstructions of cells and axons suitable for easily determining the orientation and polarity of a specimen. It also enables video reconstructions to be created, even of specimens parallel to the plastic base of a tissue culture dish, It is also helpful for studying the distribution and organization of living cells in a culture, as it provides the same powerful information as optical tomography, which most confocal microscopes cannot do on sterile living cells.