Isolation and analysis of single cells from the mouse heart

•Protocol enables preparation of cardiac non-cardiomyocyte single cell suspensions.•Protocol preserves cell surface markers, enabling immunophenotyping.•Protocol enables enrichment of specific cardiac cell populations.•Enriched cells can be used for downstream cellular and molecular analyses.•Standard or specialised laboratory equipment can be used for cell preparation.

The adult mouse heart is comprised of a highly heterogeneous cell population. Isolation and effective cellular and molecular analysis of various cell types are critical for understanding cardiac development, homeostasis and disease. Moreover, strategies to isolate and analyse the complex inflammatory and tissue remodelling cell types that follow cardiac injury are particularly important for development of strategies to improve cardiac repair. Here we describe in detail how non-cardiomyocytes can be successfully isolated from the mouse heart. In addition, we describe how these isolation methods can be effectively coupled with flow cytometry, fluorescence activated cell sorting and/or magnetic-labelling to analyse and enrich cells for subsequent cellular or molecular analyses.