Immunofluorescence analysis of Leptin was performed using 70% confluent log phase 3T3-L1 cells differentiated with StemPro Adipogenesis Supplement (Product # A10070-01) for 5 days. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Leptin Rabbit Polyclonal Antibody (Product # PA1-051) at 2µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e shows the undifferentiated cells that have lower level of Leptin expression. Panel f represents the no primary control. The images were captured at 60X magnification.