Point counting on the Macintosh. A semiautomated image analysis technique.

Abstract

In image analysis, point counting is used to estimate three-dimensional quantitative parameters from sets of measurements made on two-dimensional images. Point counting is normally conducted either by hand only or manually through a planimeter. We developed a semiautomated, Macintosh-based method of point counting. This technique could be useful for any point counting application in which the image can be digitized. We utilized this technique to demonstrate increased vacuolation in white matter tracts of rat brains, but it could be used on many other types of tissue. Volume fractions of vacuoles within the corpus callosum of rat brains were determined by analyzing images of histologic sections. A stereologic grid was constructed using the Claris MacDraw II software. The grid was modified for optimum line density and size in Adobe Photoshop, electronically superimposed onto the images and sampled using version 1.37 of NIH Image public domain software. This technique was further automated by the creation of a macro (small program) to create the grid, overlay the grid on a predetermined image, threshold the objects of interest and count thresholded objects at intersections of the grid lines. This method is expected to significantly reduce the amount of time required to conduct point counting and to improve the consistency of counts.