Spectrum plus information management system

For relative CD31 þ blood vessel density, at least 10 images (20Â objective) were taken from each animal eye (two images for the central retina, four images for the mid retina, and four images for peripheral retina) using a Zeiss Axio Imager Upright microscope. The relative CD31 þ blood vessel density in the entire image (13.5 Â 10 cm) was measured by determining the frequency of positive immunolabeled CD31 þ vascular structures coinciding with the intersection of 100 intersection points with a diameter of 0.8 cm in a 13.5 Â 10-cm frame superimposed over the image, as reported previously. 16, 17 Trypsin Digest Preparation and Quantitative Analysis of Relative Acellular Capillary Density Trypsin digests of retinal whole mounts were prepared to permit examination of the frequency of acellular capillary density using the method described by Kuwabara and Cogan, 18 with minor modifications. Retinas were scanned using the Aperio CS slide scanning system with Spectrum Plus information management system (Aperio Technologies Inc., Vista, CA). A total of 10 to 15 random, nonoverlapping fields of retina were imaged (100,000 mm 2 ). Acellular capillaries >50 mm in length were counted from each image for each retina and expressed relative to control.