Figure 4.

Despite the elevated levels of p27^Kip, less p27^Kip is bound to cdk2 in CML CD34⁺ cells. Proteins were extracted from 5–10 × 10⁶ FN-A, FN-NA, PLL-A (not shown), and PLL-NA (not shown) CML or NL CD34⁺ cells. cdk2 was immunoprecipitated from 100 μg of protein by using anti-cdk2 antibodies, or control IgG. To determine the amount of cdk2 present, the immune complexes were separated by SDS/PAGE and blots were probed with anti-cdk2 antibodies and goat anti-mouse HRP-conjugated antibody. To determine the amount of p27^Kip bound to cdk2, blots were stripped and reprobed with anti-p27^Kip and goat anti-mouse HRP antibodies. A representative example of three experiments is shown.