Bif-1 haploinsufficiency results in an increase in mitochondrial mass at premalignant stage of Myc-induced lymphoma. Prelymphomatous spleens were isolated from male littermate mice at the age of 12 weeks (panels A-C and E-F) and 10 weeks old (panel D). (A) Tissues were divided into 2 parts and a portion processed for electron microscopic analysis was stained with methylene blue and Azure II. (B) Splenocytes were prepared from another half portion of tissue, stained with MitoTracker Green FM for 30 minutes, and analyzed by flow cytometry. The percentages of cells with increased numbers of mitochondria by Myc are shown. (C-D) Geometric means of MitoTracker Green FM signals quantified and shown as relative intensities. (E) Tissue blocks in panel A were sectioned and analyzed by electron microscopy. The scale bars represent 1 μm. (F) Total mitochondrial area per cytoplasmic area in Eμ-Myc/Bif-1^+/+ (n = 136) and Eμ-Myc/Bif-1^+/− (n = 129) cells in panel E was quantified using ImageJ software and is shown as a box plot. The lines, boxes, and error bars represent median values, 25-75 percentiles, and 10-90 percentiles, respectively. Statistical significance was determined using a Wilcoxon rank-sum test with continuity correction. *P < .0001.