The potential of pluripotent embryonic stem cells (ESCs) isolated from the inner mass of blastocysts are investigated for its ability to differentiate on biocompatible electrospun nanofibers, for regeneration of the myocardially infracted heart. Nanostructured poly(d,l-lactide-co-glycolide)/collagen (PLGA/Col) scaffolds with fiber diameters in the range of 300 +/- 65 nm, was fabricated by electrospinning to mimic the extracellular matrix of the native tissue. During the culture of embryoid bodies outgrowth on the scaffolds, and further differentiation of ESCs to cardiomyocytes, the PLGA/Col nanofibers was found better than that of the electrospun PLGA nanofibers, where a better interaction and growth of ESC differentiated cardiomyocytes was observed on the composite scaffolds. The phenotypical characteristics of ESC-derived cardiomyocytes and molecular protein expression were carried out by scanning electron microscopy and immunocytochemistry, respectively. Our studies highlight the significance of a suitable material, its architecture, and cell-biomaterial interactions that is essential at a nanoscale level signifying the application of a bioengineered cardiac graft for stem cell differentiation and transplantation, which could be an intriguing strategy for cardiac regeneration. (c) 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 102B: 447-454, 2014.