Isolation of Live Label-Retaining Cells and Cells Undergoing Asymmetric Cell Division via Nonrandom Chromosomal Cosegregation from Human Cancers

The ability to retain DNA labels over time is a property proposed to be associated with adult stem cells. Recently, label retaining cells (LRC) were indentified in cancer. LRC were suggested to be the result of either slow-cycling or asymmetric-cell-division with nonrandom-chromosomal-cosegregation (ACD-NRCC). ACD-NRCC is proposed to segregate the older template DNA strands into daughter stem cells and newly synthesized DNA into daughter cells destined for differentiation. The existence of cells undergoing ACD-NRCC and the stem-like nature of LRC remain controversial. Currently, to detect LRC and ACD-NRCC, cells need to undergo fixation. Therefore, testing the stem-cell nature and other functional traits of LRC and cells undergoing ACD-NRCC has been limited. Here, we show a method for labeling DNA with single and dual-color nucleotides in live human liver cancer cells avoiding the need for fixation. We describe a novel methodology for both the isolation of live LRC and cells undergoing ACD-NRCC via fluorescence-activated cell sorting with confocal microscopy validation. This has the potential to be a powerful adjunct to stem-cell and cancer research.