Adhesive Signature-Based, Label-Free Isolation of Human Pluripotent Stem Cells

Generation of human induced pluripotent stem cells (hiPSCs) from fibroblasts and other somatic cells represents a highly promising strategy to produce auto- and allo-genic cell sources for therapeutic approaches as well as novel models of human development and disease¹. Reprogramming protocols involve transduction of the Yamanaka factors Oct3/4, Sox2, Klf4, and c-Myc into the parental somatic cells, followed by culturing the transduced cells on mouse embryonic fibroblast (MEF) or human fibroblast feeder layers, and subsequent mechanical dissociation of pluripotent cell-like colonies for propagation on feeder layers^{1, 2}. The presence of residual parental and feeder-layer cells introduces experimental variability, pathogenic contamination, and promotes immunogenicity³. Similar to human embryonic stem cells (hESCs), reprogrammed hiPSCs suffer from the unavoidable problem of spontaneous differentiation due to sub-optimal feeder cultures⁴, growth factors⁵, and the feeder-free substrate⁶. Spontaneously differentiated (SD)-hiPSCs display reduced pluripotency and often contaminate hiPSC cultures, resulting in overgrowth of cultures and compromising the quality of residual pluripotent stem cells⁵. Therefore, the ability to rapidly and efficiently isolate undifferentiated hiPSCs from the parental somatic cells, feeder-layer cells, and spontaneously differentiated cells is a crucial step that remains a bottleneck in all human pluripotent stem cell research.