Novel Cell Surface Marker for the Identification of iPSC in Somatic Cell Reprogramming

The creation of human induced pluripotent stem cells (iPSCs) and their downstream applications demonstrate great promise as valuable tools for disease research, therapeutic applications and biological engineering. As different somatic tissues are reprogrammed into a pluripotent state using different technologies and under varying culture conditions, the critical step of establishing iPSC lines requires the early identification of true iPSC clones. It is important to identify and pick the right colonies for further expansion and characterization.  Commonly, surface antibodies against pluripotent specific markers such as SSEA4, Tra-1-60 and Tra-1-81 are used but this method is expensive and sterility is a concern.  We have utilized a global transcriptome comparison of human parental fibroblasts, and partially and fully reprogrammed clones to identify differentially expressed markers. One such marker is the cell adhesion surface molecule, CD44, which is highly expressed in parental fibroblasts and partially reprogrammed cells, but not in fully reprogrammed iPSCs.

CD44 has been used in combination with pluripotent markers to show distinct expression patterns between fully reprogrammed pluripotent colonies and partially or non-reprogrammed colonies during reprogramming. In this study, we utilized antibodies against CD44, in conjunction with other known pluripotent surface markers for elucidating reprogramming kinetics and for the rapid enrichment of pluripotent stem cells from reprogrammed somatic tissues. This combined approach of positive and negative markers for identification of true iPSCs during the reprogramming process permits the selection of bonafide clones during cell line creation.