Effect of detergent concentration on GEM preparation. WEHI-231 cells were left unstimulated (–) or were stimulated (+) with 20 μg/mL anti-IgM Ab for 5 minutes. Cells were lysed in 0.5%, 0.25%, and 0.1% Triton X-100 and were subjected to sucrose gradient ultracentrifugation. Fractions 4 and 5 combined were then separated by SDS-PAGE and blotted with anti-CD45 mAb, anti-CD71 mAb, and HRP-conjugated CTB subunit to detect GM-1.