mmunofluorescence analysis of Vimentin was performed using 70% confluent log phase HeLa cells. 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 Vimentin Mouse Monoclonal Antibody (Product # MA1-19168) at 5 µg/mL in 0.1% BSA and incubated overnight at 4 degree Celsius and then labeled with Goat anti-Mouse IgG (H+L)/IgM (L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) 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, cytoskeletal and nuclear localization. Panel e represents negative control, A-431 cells. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.