Protocol for generating human iPSC-derived cardiomyocytes using VCAM1-based sorting and micropatterned culture

The expanding use of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) in disease modeling and regenerative medicine has highlighted persistent challenges, including inter-differentiation variability, immature phenotypes, and heterogeneous cell populations. Here, we present a protocol for generating human iPSC-CMs by combining fluorescence-activated cell sorting (FACS) of VCAM1-positive cells with culture on micropatterned substrates. We describe steps for inducing cardiac differentiation in 3D and enriching iPSC-CMs by FACS. We then detail procedures for maturing iPSC-CMs in 2D micropatterned culture. For complete details on the use and execution of this protocol, please refer to Gomes-Silva et al.¹.