Expression of Akt and p27 in ALCL primary tumors. (A) Subcellular fractionation followed by Western blot analysis demonstrated that Akt and pAkt are predominantly localized in the cytoplasm of Karpas 299 and SU-DHL1 cells. Retinoblastoma protein (Rb) was detected exclusively in the nuclear fraction and served as a control for nuclear localization of proteins in these cells. (B) Box-plot showing the significant difference in the percentage of p27-positive tumor cells between pAkt-positive and pAkt-negative ALCL tumors (P = .0076). Only nuclear expression of total p27 was considered positive in this analysis. Error bars indicate 5%-95%. (C-D) Immunohistochemical detection of pAkt in ALCL tumors showed strong cytoplasmic expression of pAkt in a subset of ALK-positive and ALK-negative ALCL tumors, shown in panels C and D, respectively. (Immunohistochemical staining, 600 × original magnification, obtained with a BX51 Olympus microscope [Melville, NY] and a DP12 Olympus camera.) Images were viewed through a universal semi-apochromat objective lens (UPlan Fl, Olympus America Inc, Woodbury, NY); original magnification × 400; numerical aperture 0.75.