Computer-assisted cytophotometry is a technique used to quantitate the size, amount and distribution of specifically-stained material within cells. When combined with image analysis techniques, computer-assisted cytophotometry becomes a very powerful tool for biomedical research. Current cytophotometers are relatively expensive, often slow, and many of the commercial instruments do not have the flexibility required for biomedical research. During the past decade charge-coupled device (CCD) imaging technology has developed. These new solid-state devices feature high speed, high sensitivity, low noise and a fixed geometry that makes this new technology attractive for biomedical research applications using cytophotometry. This New Investigator Research Award application presents a proposal to develop, test and apply a computer-assisted cytophotometry system. This new system will consist of a Fairchild CCD image camera, a Zeiss Universal microscope, and a Digital Equipment Corporation LSI 11/23 computer. Computer programs will be developed for acquisition of data from the CCD device, image segmentation, image display, and calculation of total optical density, cell area, and shape features. Precision and accuracy of the measurements will be made by comparing results obtained with the CCD camera with measurements on the same cells made with a Zeiss scanning stage cytophotometer using the HIDACSYS computer programs. The system will then be applied to both clinical and basic research projects, including: testing the effects of immunostimulants on the lysosomal enzyme content of macrophages cultured in tissue culture; rapid detection of enzyme differences in polymorphonuclear neutrophils (PMNs) from septicemic infants; acquisition of cell images for monitoring patients with chronic lymphocytic leukemia; real-time detection of changes in fluorescence intensity for the study of the mechanisms of host resistance to intracellular pathogens; and measurement of direction and speed of leukocyte movement in tissue culture. Development of a CCD-based computer-assisted cytophotometry system will benefit future biomedical research by providing a lower cost, faster and more versatile instrument.
