A New Approach to Assess Viability of Adult Cardiomyocytes: Computer-Assisted Image Analysis

Objective:to develop a computerized procedure to obtain the percentage of rod-shaped cells in preparations of isolated adult cardiomyocytes.Methods:(1) isolated adult rat myocytes were either pretreated with a fixative (0.2% glutaraldehyde, 0.25% Triton X-100 and 0.1% trypan blue) or monitored via phase-contrast optics without fixation; (2) an image field with several hundred cells was captured by a microscope-mounted video camera, which was connected to a frame grabber, and saved as a TIFF file; (3) the image was processed using Mocha software. Analysis consisted of setting the image threshold, an object count, automatic measurements of objects and their classification.Results:though the software could measure many geometrical characteristics, a combination of two parameters, object size and shape factor, was found to be the most efficient. With these parameters, four classes of sample objects were constructed including rod-shaped myocytes, round damaged cells, overlapping cell clusters, and small debris. Test objects were classified automatically based on which sample object their parameters matched the most. Excellent correlation was obtained between manual counts and computer analysis of cell viability (r=0.98).Conclusion:a more efficient method, based on computer-assisted classification of objects, has been developed to assess the viability of isolated adult cardiomyocytes.