A SIMPLE METHOD FOR CALIBRATING COLLAGENASE PRONASE-E RATIO TO OPTIMIZE HEART CELL ISOLATION/

A mixture of crude collagenase and non-specific proteases has been use d to isolate guinea pig ventricular heart cells. Measurements of colla genase activity with Wunsch's substrate and protein content with sodiu m dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) sugges t that collagenase enzymes do not play a major role in heart cell isol ation. On the other hand, an important factor in heart digestion seems to consist of some fractions of the proteases present in crude collag enase. It is also noted that crude collagenases do not present any sen sitivity to added calcium but because this ion is important to obtain isolated cells its role is discussed. According to our results, the SD S-PAGE method can be used to determine the appropriate enzyme concentr ations to obtain calcium-tolerant myocytes. These myocytes have electr ophysiological properties as reported in the literature.