Immunofluorescence analysis of TGF beta1 was performed using THP-1 and THP-1 differentiated and polarized to M2 macrophage cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with TGF beta-1 Polyclonal Antibody (Product # PA1-29032) at 1:100 in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790), (1:2500 dilution), for 45 minutes at room temperature (Panel a, e: Green). Nuclei (Panel b:Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing increased expression and cytoplasmic localization of TGF beta1 in M2 macrophage compared to control THP-1 cells (Panel e) . Panel f represents control M2 macrophage cells with no primary antibody to assess background. The images were captured at 60X magnification.