Human pluripotent stem cells, such as induced pluripotent stem cells (hiPSCs) and human embryonic stem cells (hESCs), hold significant promise for advancing both basic cell research and clinical applications. A major hurdle in harnessing their full potential is the efficient and accurate selection of clones that are both functional and pure. Traditional methods, such as manual colony picking and limiting dilution, are either low-throughput, labor-intensive or lack the precision required for selecting the most suitable clones. These methods also struggle to maintain the pluripotency of colonies, which is crucial for applications requiring cell differentiation, such as disease modeling. Identifying and selecting stem cells that remain undifferentiated is possible using fluorescence-activated cell sorting (FACS) to isolate cells based on the expression of specific markers, such as SSEA-4 and TRA-1-60-R. For efficient single-cell sorting and cultivation, technologies like the WOLF G2® Cell Sorter and the N1 Single-Cell Dispenser have shown promise. These tools can gently deposit a single cell into a well using low pressure (less than 2 psi), which supports the growth of undifferentiated colonies at a higher rate than traditional methods like limiting dilution. We have successfully utilized the WOLF G2 to sort GM23338 stem cells with over 90% efficiency. This process resulted in more than 40% clonal outgrowth, with the cells maintaining expression of key pluripotency markers SSEA-4 and TRA-1-60-R, indicating a significant improvement in the efficiency and reliability of stem cell cloning techniques.
PST-017
