Fig. 1.
(a) HPTLC analysis of plasma membrane phospholipids in anti-Fas-treated U937 cells. Apoptosis was induced by incubating the cells (5 × 105 cells/ml), with anti-Fas (CD95) IgM mAb, 100 ng/ml for 4 h. Plasma membrane fractions from U937 cells were isolated by continuous 25–65% (w/v) density gradients of sucrose. Phospholipids were extracted according to the technique described by Folch [22] and separated by thin-layer chromatography, in chloroform: methanol: acetic acid: water (100:75:7:4) (v:v:v:v). Phospholipids were stained with iodide vapours. Lane A, phospholipids obtained from untreated U937 cells; lane B, phospholipids obtained from anti-Fas treated U937 cells. (b) Immunoblotting analysis of the cytochrome c (mitochondrial marker) contamination in plasma membrane preparation. Whole cell extract and the plasma membrane preparation were subjected to SDS-PAGE (12%), Western blot and immunodetection of cytocrome c using a specific monoclonal antibody. CN, whole cell extract; PM, plasma membrane preparation.