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. Author manuscript; available in PMC: 2010 May 3.
Published in final edited form as: J Biol Rhythms. 2008 Feb;23(1):26–36. doi: 10.1177/0748730407311254

Figure 2.

Figure 2

BMAL1 expression oscillates in Leydig cells. (A) Immunohistochemical analysis of sectioned testes detected BMAL1 almost exclusively in the Leydig cells. Top panel: Merged images of BMAL1 (green) and nuclear dye Hoechst 33258 (blue). Bottom panel: Staining for BMAL1 only. BMAL1 staining intensity oscillates over the course of day, with a peak at ZT3 and a trough at ZT9. Images shown are representative of 5 testes processed at each time point. (B) Preadsorption of antibody with BMAL1 peptide dramatically reduces staining. Magnification in all pictures is ×40. (C) BMAL1 oscillates in both cytoplasm (cyt) and nuclei (nuc), as measured by densitometry. Data represent the average (± SEM) level of staining in 20 sections derived from 5 animals per time point. The vertical axis represents relative staining intensity, where 0 represents no staining (black) and 255 represents maximum saturated staining (green) assigned by ImageJ software. Significant differences (p < 0.05) in BMAL1 signal in both cytoplasmic and nuclear staining were observed between all time points, except between ZT21 and ZT15 (paired Student t test). Minimal staining using antibody preadsorbed with peptide was observed in cytoplasm (cyt-PR) and nuclei (nuc-PR).