Differential expression of p27 and miR-221/222 in different CLL tissue compartments. Western blot analysis with the use of whole-cell lysates of purified CLL B cells from paired LN/BM and PB obtained from 7 patients. The blots were probed with anti-p27 antibodies; β-actin was used as control for equal protein loading. A representative case is shown (A top). The differences in p27 expression are quantified by densitometric analysis (A bottom). Quantitative PCR analysis (B) for miR-221 and miR-222 (TaqMan miRNA assay) quantifies the differences in expression between purified leukemic cells from paired LN/BM and PB from the same 7 patients depicted in panel A. Statistical significant differences were evaluated by the Wilcoxon test. Immunofluorescent staining with p27 (green), propidium iodide (PI; red), and Ki67 (blue) of CLL cells purified from LNs (first and third columns, 63× and 240× magnification, respectively) and PB cells (second and fourth columns, 63× and 240× magnification, respectively) of a representative patient with CLL (C; 63×/1.25 NA oil objective). LN cells exhibit lower levels of p27 in comparison to PB cells. Ki67 staining could detect positive cells only in LN cells, whereas PB cells are negative.