Fig. 2.
mAb-mediated stimulation of hCD28 in Jurkat T cells leads to its translocation to lipid rafts. Jurkat cells (108 cells per ml) were incubated with medium alone or with anti-CD28 mAb for 5 min at 37°C. The lysates of the cells were then prepared for sucrose centrifugation according to Materials and Methods. CD28 immunoprecipitates of the 12 fractions were analyzed by immunoblotting with antiphosphotyrosine (PY), anti-CD28, and anti-PI3K p85. Each fraction was also subjected to blotting with anti-Lck, anti-actin, and HRP-conjugated cholera toxin B, which marks the presence of GM1.