Isolation and culture of neonatal mouse cardiomyocytes.
Isolation and culture of neonatal mouse cardiomyocytes.
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DOI：
10.3791/50154
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发表时间：
2013-09-06
期刊：
影响因子：
--
通讯作者：
Lange, Stephan
中科院分区：
文献类型：
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作者：
Ehler, Elisabeth;Moore-Morris, Thomas;Lange, Stephan
长期以来，培养的新生心肌细胞被用于研究肌原纤维发生和肌原纤维功能。培养的心肌细胞允许容易的调查和操作的生化途径，以及它们对自发跳动的心肌细胞的生物力学特性的影响。以下2天方案描述了新生小鼠心肌细胞的分离和培养。我们展示了如何容易地从新生儿中解剖心脏，分离心脏组织并从心肌细胞群中富集心肌细胞。我们讨论了使用不同的酶混合物的细胞解离，以及它们对细胞活力的影响。分离的心肌细胞随后可用于各种形态学、电生理学、生物化学、细胞生物学或生物力学测定。我们通过仅使用市售溶液和酶混合物优化了方案的耐用性和重现性，这些溶液和酶混合物显示出很小的批间变异性。我们还解决了与心肌细胞的分离和培养相关的常见问题，并提供了各种优化分离和培养条件的选择。
Cultured neonatal cardiomyocytes have long been used to study myofibrillogenesis and myofibrillar functions. Cultured cardiomyocytes allow for easy investigation and manipulation of biochemical pathways, and their effect on the biomechanical properties of spontaneously beating cardiomyocytes. The following 2-day protocol describes the isolation and culture of neonatal mouse cardiomyocytes. We show how to easily dissect hearts from neonates, dissociate the cardiac tissue and enrich cardiomyocytes from the cardiac cell-population. We discuss the usage of different enzyme mixes for cell-dissociation, and their effects on cell-viability. The isolated cardiomyocytes can be subsequently used for a variety of morphological, electrophysiological, biochemical, cell-biological or biomechanical assays. We optimized the protocol for robustness and reproducibility, by using only commercially available solutions and enzyme mixes that show little lot-to-lot variability. We also address common problems associated with the isolation and culture of cardiomyocytes, and offer a variety of options for the optimization of isolation and culture conditions.