Methods of mouse cardiomyocyte isolation from postnatal heart

Primary cardiomyocytes (CMs) are invaluable Materials used to study in vitro cardiac physiology and pathophysiology. However, there is still a lack of comprehensive, systematic and comparative studies on methods for isolation of primary CMs from postnatal hearts. Here, we aimed to compare and optimize protocols for the isolation of CMs from embryonic to adult stages. We found that the trypsin digestion method was suitable for embryonic mouse CM isolation. The Gentle MACS method yielded high-quality CMs from neonatal hearts (postnatal day 1-day 3, P1-P3). The Langendorff-free perfusion method was applicable for isolation of CMs from mice older than P3. P14 and P56 CMs could also be isolated by the Langendorff perfusion system. The transcriptional profiles and cellular function of the isolated CMs were respectively confirmed by RNA sequencing and Angiotensin II treatment, suggesting the feasibility and effectiveness of the isolation methods. Overall, this study supplies a series of detailed and optimized protocols to isolate CMs at different developmental stages, which provides support for the study of the mechanisms, pathology, and pharmacology of cardiovascular diseases.