There is a lack of organs and tissues for patients in need. Human pluripotent stem cells (hPSCs), which encompass human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), have been considered to be promising sources for regenerating damaged tissues and organs because of their ability to differentiate into cells from three embryonic germ layers. Currently, only for four main diseases are targeted for the clinical trials for stem cell therapy using hPSCs: (1) macular degeneration [age-related macular degeneration (AMD) and Stargardt’s macular dystrophy], (2) acute myocardial infarction (MI), (3) diabetes, and (4) spinal cord injury. The efficient and reproducible differentiation method of hPSCs into cardiomyocytes has not yet investigated from our database research. Especially, it is unclear what kinds of molecular, physical, and biological design of cell culture biomaterials are preferable for differentiation and purification (sorting) of hPSCs into cardiac progenitors and cardiomyocytes. Therefore, the development of sophisticated smart biomaterials, which are optimized for the differentiation and sorting of hPSCs into cardiomyocytes and cardiac progenitors are important for future stem cell therapies. In this study, we are going to develop optimal cell culture smart biomaterials for differentiation and sorting of hPSCs into cardiac progenitors and cardiomyocytes in xeno-free conditions. We will investigate (i) what type of smart biomaterials are desirable for differentiation of hPSCs into cardiomyocyte progenitors and cardiomyocytes and for sorting of hPSC-derived cardiomyocyte progenitors and cardiomyocytes, and (ii) what kind of nanosegments or extracellular matrices (ECMs) should be immobilized on cell culture biomaterials coated with thermoresponsive polymer (poly(N-isopropylacrylamide), PNIPAAM copolymer) for culture and differentiation of hPSCs as well as sorting of hPSC-derived cardiac progenitors and cardiomyocytes on smart biomaterials. Our originality of the smart biomaterial development for stem cell culture, differentiation, and sorting is the combination of physical cues (thermoresponsive polymer-coated dishes to detach specific cells by reducing temperature of the culture medium) and biological cues (selection of specific extracellular matrix (ECM) or ECM-derived oligopeptide for recognition of cardiac progenitors and cardiomyocytes). We are going to develop cell culture smart biomaterials immobilized with thermoresponsive polymer (PNIPAAM copolymer), which are further coatd with ECM or ECM-derived oligopeptides, which are suitable for stem cell culture, differentiation, and sorting into cardiac progenitors and cardiomyocytes in clinical usage in future. PNIPAAM copolymer is selected as the thermoresponsive polymer for the coating on the cell culture dishes, which has low critical solution temperature (LCST) around 20 degree. Several ECM proteins and oligopeptides are further immobilized on the PNIPAAM copolymer. The hiPSCs are cultured on the thermoresponsive dishes grafted with ECM and oligopeptides and are differentiated into cardiac progenitors and cardiomyocytes using several differentiation protocols from articles and from our developed method in this study. We evaluate the efficiency and purity of cardiomyocyte differentiation from the data and discuss the optimal cell culture smart biomaterials for specific differentiation and sorting of hPSCs into cardiac progenitors and cardiomyocytes.