HCA7 cells were seeded at a density of 0.5 Â 10 6 cells per well in six-well plates and were treated with SC236 (5 mM) or vehicle control for 24 h. Cells were detached using Versene for 5 min at 371C and floating and adherent cells were then pelleted by centrifugation and washed in a solution of 2% BSA suspended in PBS. We performed FITC -Annexin V/propidium iodide labelling was performed using a TACS Annexin V -FITC Apoptosis detection kit (R&D Systems, Abingdon, UK). Briefly, cells were re-suspended in a 100 ml working stock of binding buffer, propidium iodide and Annexin V -FITC conjugate and incubated in the dark for 15 min. Cells were re-suspended in binding buffer before analysis in a FACScalibur flow cytometer (Becton Dickinson, Oxford, UK) with measurement of fluorescence emission at 530 nm (FL1 channel) and at 4575 nm (FL3 channel). Four quadrant analyses using CellQuest (Becton Dickinson, Oxford, UK) software allowed quantification of cell populations according to labelling characteristics. The results were verified by staining/morphology by confocal microscopy.