Bif-1 haploinsufficiency up-regulates Mcl-1 expression and suppresses caspase-3 activation in Eμ-Myc–induced tumors. Lymphomatous axillary lymph nodes were extirpated from each Eμ-Myc–transgenic mouse at the onset of disease. (A) Tissue lysates were prepared using a Tissuemiser homogenizer and analyzed by immunoblotting using the indicated antibodies. Representative blots of Eμ-Myc/Bif-1^+/+ (n = 37), Eμ-Myc/Bif-1^+/− (n = 38), and Eμ-Myc/Bif-1^−/− (n = 8) tumors in addition to control Bif-1^+/+ and Bif-1^+/− lymph nodes are shown. (B) The intensity of cleaved-Caspase-3 (C-Casp-3, p17), Mcl-1, and p62 were quantified by densitometry using Quantity One software and are shown as a box plot. To compare the signal intensities between the different gels, the intensity of each blot was adjusted to the value of untreated wild-type mouse embryonic fibroblast lysates, which were loaded on every gel. Statistical significance was determined using 2-way ANOVA followed by the Scheffe posthoc test. *P < .05. (C-D) Tissues subjected to immunohistochemical analyses using the indicated antibodies. The number of proliferating cell nuclear antigen (PCNA)–positive cells per field (400×) was counted using a light microscope and is shown as a box plot in panel D. A total of 7451, 7251, and 9947 PCNA⁺ cells were counted from 3 Eμ-Myc/Bif-1^+/+, 3 Eμ-Myc/Bif-1^+/−, and 4 Eμ-Myc/Bif-1^−/− mice, respectively.