Immunofluorescence analysis of Cytokeratin 18 was performed using 70% confluent log phase Caco-2 cells. The cells were fixed with ice-cold acetone at 4°C for 5 minutes and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with Cytokeratin 18 Monoclonal Antibody (DA-7) (Product # MA1-19047) at a concentration 5 µg/mL in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175), (1:2000 dilution), 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 dilution). Panel d represents the merged image showing cytoskeletal localization. Panel e represents A-375 cells showing no cytoskeletal staining. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.