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. 2014 Jun 27;5(13):4694–4708. doi: 10.18632/oncotarget.2100

Figure 5. Expression of cell cycle regulators and cyclin-Cdk-p27 complexes in CLL cells.

Figure 5

(A) Representative immunoblot showing the levels of cyclins A and E in CLL cells with and without Myc/p27. Actin levels were also determined to assess protein loading. (B) Correlation between Myc and cyclin A protein levels in CLL cells. (C) Immunoblot showing Myc and p27 protein levels in four patient's cells selected for the experiments shown in D. (D) Molecular filtration chromatographic separation of CLL protein extracts followed by immunoblot for cyclins A, E and Cdk2. The elution of the 55 and 30 kDa proteins is shown at the top. (E) Immunoblot (IB) showing the levels of Myc and p27 in three CLL samples and MEC1 cells and kinase assay of Cdk2 in the same extracts. Proteins were immunoprecipitated with anti Cdk2 and the presence of both cyclin E, p27 and Cdk2 were determined by immunoblot. Lower panel: kinase assays were performed using HisCK− as kinase substrate. M, mock kinase reaction without extract. (F) Immunoprecipitation of Cdk2 and kinase assay of the immunoprecipitates of MEC1 cells and MEC1 cells incubated with lysates from a CLL sample (#01). (G) Immunoprecipitation of Cdk2 and kinase assay from MEC1 cells and from mixed lysates prepared with MEC1 cells and two CLL samples (#01 and #10).