After dissociating both populations of cells by Tripsin-EDTA, check to see if there are notable differences in size or density. If there are, an isopicnic gradient ultracentrifugation would do the job (done with Percoll-HBS). Cells are enriched in the appropriate density band. - If you can get neutralizing antibodies against voltage-gated calcium or sodium channels or any other surface molecule in the cardiomyocyte, you might try the magnetic beads by either positive (you keep your eluate) or negative selection (you keep the filtrate). - As you suggest, FACS is another choice, expensive in my view, because you have to get primary antibodies against the same targets conjugated to fluorochromes, plus service fees, the day becomes hectic. Another option is to kill the cells you don't want (Neutralizing antibody, addition of complement).

Label fibroblasts with PKH26 and cardiomycocytes with PKH67 and co-culture. The cells can be then sorted by FACS. These dyes stay in cells for minimum a week. See Figure 3A of my paper (cancer cells+immune cells)

Paper: The Blebbishield Emergency Program Overrides Chromosomal Instability and Phagocytosis Checkpoints in Cancer Stem Cells | Cancer Research (aacrjournals.org)

Hi Giorgia, I don't know if your experiment requires direct or indirect co-culture. If direct, you will need to label your cells and then do FACS.

If it is indirect co-culture, then it is easier. You can use cell inserts (cellQART for example). You can grow fibroblasts on the bottom of the plate, and cardiomyocytes on the cell insert. That way you have both cells in the same well, but without direct contact. Hope it helps! Good luck!

If you are using adult cardiac myocytes and if they are attached to laminin-coated dishes, they require longer time incubation with trypsin. So you can detach Fb within few minutes of incubation and later you can harvest cardiac myocytes.