Isolation, culture, and immunostaining of neonatal rat ventricular myocytes

ABSTRACT: Summary Isolation and culture of ventricular cardiomyocytes from neonatal rats (NRVMs) is a powerful model to study neonatal cardiac development, cell cycle regulation, and cardiac physiology and pathology in vitro. Here, we present our modified enzymatic digestion protocol followed by two-step discontinuous Percoll gradient centrifugation to isolate a high yield of viable ventricular cardiomyocytes from neonatal rats. Finally, here we describe an immunostaining protocol for cytosolic and nuclear staining of NRVMs. For complete details on the use and execution of this protocol, please refer to Pereira et al. (2020). Graphical abstract Highlights • Isolate a high yield of viable ventricular cardiomyocytes from neonatal rats (NRVMs)• NRVMs can be maintained in culture for several days• Enzymatic digestion of cardiac tissue and Percoll gradient separation• Detailed subsection of immunofluorescence for nuclear and cytosolic staining Isolation and culture of ventricular cardiomyocytes from neonatal rats (NRVMs) is a powerful model to study neonatal cardiac development, cell cycle regulation, and cardiac physiology and pathology in vitro. Here, we present our modified enzymatic digestion protocol followed by two-step discontinuous Percoll gradient centrifugation to isolate a high yield of viable ventricular cardiomyocytes from neonatal rats. Finally, here we describe an immunostaining protocol for cytosolic and nuclear staining of NRVMs.

SUBMITTER: Pereira A 

PROVIDER: S-EPMC8599495 | biostudies-literature |

REPOSITORIES: biostudies-literature