Figure 1

Analysis of Aurora-A expression by western blot, immunocytochemical, and immunohistochemical analyses. (a) Western blot analysis was performed using CD19+ B-cell lysates of chronic lymphocytic leukemia (CLL) and control subjects. MCF-7 breast cancer cells were used as a positive control. The graph shows the distribution of patients along the median in terms of Aurora-A levels expressed as the ratio of Aurora-A to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). (b) Immunohistochemical analysis in a case of typical CLL involving bone marrow. Aurora-A expression was both nuclear and cytoplasmic and was more prominent in the larger prolymphocytes and paraimmunoblasts than in the smaller neoplastic cells. Double-labeling immunohistochemical staining with Aurora-A (red) and the B-cell marker anti-CD20 (brown) showed coexpression of Aurora-A in the CD20+ neoplastic B-CLL cells (inset). (c) Immunohistochemical analysis in a case of staging bone marrow, with a benign lymphoid aggregate used as a control for Aurora-A expression. Aurora-A was expressed in rare lymphocytes, especially in immunoblasts, whereas the smaller benign lymphoid cells were negative (inset). (d) Immunocytochemical analysis of subcellular localization of Aurora-A in CLL cells. Formalin-fixed bone marrow smears of untreated patients with CLL were used for immunocytochemical analysis. Aurora-A was expressed in both the cytoplasm and the nuclei of most of the tumor cells. (e) Immunocytochemical analysis of subcellular localization of phosphorylated Aurora-A in CLL cells. The phosphorylated form of Aurora-A and total Aurora-A were distributed similarly, both within the cytoplasm and nuclei of tumor cells. Compared with total Aurora-A, the phosphorylated form was expressed in fewer tumor cells. A benign lymphocyte, negative for phosphorylated Aurora-A, is present in the smear.