Enrichment of cardiomyocytes from neonatal rat ventricular tissues by discontinuous percoll gradient centrifugation

Objective To find a method for enrichment of cardiomyocytes from neonatal rat ventricular tissues.Methods Neonatal rat ventricular tissues were digested by trypsin.After discontinuous percoll gradient centrifugation,each cell layer was examined by light microscopy and immunohistochemistry.Cardiomyocytes were identified by immunohistochemistry and Confocal Laser Scanning Microscopy (CLSM).Results After discontinuous percoll gradient centrifugation,6 cell layers were formed.The majority of cardiomyocytes were identified in the fourth and fifth layers by light microscopy and immunohistochemistry.These cardiomyocytes spread over in the first day with individual cells beating spontaneously.On the day 3,the cells were arranged in loose and interlacing networks of single cells.High-power CLSM demonstrated that cell bundles mainly consisted of cardiomyocytes with sarcomeres in registry.These cells beat spontaneously and remained in good morphology and vitality 2 or 3 months later.Conclusion Cardiomyocytes disassociated from neonatal rat ventricular tissues can be enriched by method of discontinuous percoll gradient centrifugation.