Fig. 4.

Analysis of digestion efficiency, cell viability and the CSP profile using manual dissociation techniques. Following manual dissociation protocols the efficiency of the digestion was assessed by establishing a “singlet” gate and the number of cell doublets and aggregates quantified. For CSP analysis, following digestion and Hoechst labelling (5ug/ml), single cells were gated based on their properties displayed on the Hoechst blue area (3-355/405/50-A) versus height (3-355/405/50-H) dot plot. CSP cells are recognized as a dim tail extending first on the left side of the main population of cells toward the lower “Hoechst Blue” signal (SP gate). The Hoechst Blue vs Hoechst red plots were also used to assess the ability of FTC (10 μM) to inhibit the CSP (as identified by an absence of events in the SP gate) and the effects on cell viability. Viable cells are defined by exclusion of dead cells as recognized by their strong positivity for Propidium Iodide on the 3-355/675lp-A axis. Data represents the mean±SD of a least three individual experiments