Peptide-induced apoptosis in primary CLL cells. (A) CLL cells from 8 patients were treated with 10μM of each peptide shown and the percentage of dead cells, identified by trypan blue dye uptake, was determined 24 hours later; results are presented as mean ± SE. (B) CLL cells from 4 different patients were treated with 10μM of each peptide shown, and 24 hours later the percentage of cells (mean ± SE) with typical apoptotic morphology was determined by fluorescence microscopic analysis of Hoechst-stained nuclei. (C) Images of Hoechst-stained nuclei illustrating apoptotic morphology 24 hours after treatment with 10μM TAT-IDP_DD/AA. (D) Experiments using CLL cells from 6 patients to quantify the percentage of dead cells (mean ± SE) by flow cytometric analysis of propidium iodide uptake 24 hours after treatment with 10μM of the peptides shown. (E) Flow cytometric analysis of the same samples shown in panel D quantifying the percentage of annexin V–positive cells (mean ± SE) 24 hours after treatment with 10μM of the peptides shown. (F) Caspase-3/7 activation induced by TAT-IDP_DD/AA (mean ± SE, 2 experiments). (G) Representative confocal images of CLL cells loaded with TMRM, indicating loss of mitochondrial membrane potential 24 hours after adding TAT-IDP_DD/AA. Top panels are bright-field images; bottom panels are fluorescence images. Bar = 8 μm. (H) Inhibition of TAT-IDP_DD/AA–induced CLL cell death by xestospongin C (10μM) added 30 minutes before peptides, based on analysis of 85 cells, representative of 3 experiments using CLL cells from different patients.