Detection of CD44 variant isoform expression on the CD44^highESA^high cell sub-population.

<p>FACS analysis of CD44 variant isoform expression on cells of the CA1 (A), Met1 (B) and Met2 (C) cell lines isolated using enzyme-free buffer showing the CD44 variants on the y-axis and ESA expression on the x-axis. For all cell lines, similar total levels of CD44 are detected on the CD44^highESA^low and CD44^highESA^high cells but lower levels of the v3, v5, v6 and v9 isoforms are detected on the CD44^highESA^low cells. (D) FACS analysis of CD44 (y-axis) and ESA (x-axis) expression on CA1 (top), Met1 (middle) and Met2 (bottom) cells treated with either trypsin (left) or enzyme-free buffer (right). Loss of detection of CD44 variant isoforms after trypsin treatment results in a large increase in the fraction of motile CSCs within the 5% of cells showing the highest CD44 expression. Representative plots. (E) Quantification of the experiments depicted in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057314#pone-0057314-g004" target="_blank">figure 4D</a>, showing the percentage of EMT CSCs within the 5% CD44^high population after treatment with trypsin or enzyme-free buffer.