Immunofluorescence analysis of PH-interacting protein (PHIP) was performed using 70% confluent log phase A-431 and U-87 MG 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 1 hour at room temperature. The cells were labeled with PHIP Polyclonal Antibody (Product # PA5-77167) at 1:100 in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32731), (1:2500), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with Hoechst 33342 (Product # H1399). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear localization for PHIP in A-431 and low to no signal in U-87 MG (Panel e) which is reported low expressing for the protein. Panel f represents control A-431 cells with no primary antibody to assess background. The images were captured at 40X magnification with CellInsight CX7 LZR High-Content Screening (HCS) Platform (Product # CX7C1115LZR).