Figure 7

Analysis of detergent-resistant (isolated using 1% Triton X-100) lipid rafts. Lipid rafts were isolated from un-stimulated YH16.33 T cells (B), and YH16.33 cells co-cultured with A20 cells in the absence of antigen (C-E). Rafts were captured with anti-Thy-1 on the nickel grids followed by their detection with biotinylated anti-Ly-6A.2 and anti-biotin colloidal gold. Various sizes of lipid rafts ranging from less than 100 nm to several μm were visualized (C-E) and quantified (F). Feret's diameter of lipid rafts was determined for rafts isolated from YH16.33 alone (open squares), YH16.33 with A20 cells in the absence of antigen (light square), YH16.33 with A20 in the presence of antigen (grey square) and YH16.33 with A20 exposed to MβCD (black square) are shown. Asterisk (*) indicates significant differences and n.s denotes not significant differences from the YH16.33 alone group (F). Absence of lipid rafts isolated from A20 cells alone using anti-Thy-1 and anti-Ly-6A.2 antibodies to capture and detect, respectively is shown in A (negative control). Three independent experiments were carried out and 5 photographs were taken at 5 distinct regions on each grid. bar = 100 nm.