Abstract

Human induced pluripotent stem cells (hiPSCs) and CRIPSR-Cas gene-editing are scientific breakthrough technologies. In combination, they provide a unique opportunity to mechanistically dissect disease mechanisms, develop patient-specific disease models and significantly advance drug development. However, hiPSCs are inherently sensitive cells and a critical bottleneck in the efficient generation of gene-edited hiPSCs is the generation of single-cell derived (clonal) cultures. Existing methods for single-cell isolation are often harsh on cells, manually tedious and/or do not guarantee sufficient proof of monoclonality. Here, we detail automated single-cell isolation workflows utilizing a novel type of culture chamber that is uniquely suited to easily visualise and document single cells, as well as track outgrowing cultures. Our streamlined and automated workflows facilitate a gentle isolation and culture process, producing market-leading/industry-leading hiPSC single-cell cloning efficiencies without the need for complex protocols or instrumentation.