Immunofluorescence analysis of MCM7 was performed using DU 145 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with MCM7 Polyclonal Antibody (Product # PA5-79651) at 5 µg/mL in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: 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 nuclear expression of MCM7 in DU 145. Panel e represents control DU 145 cells with no primary antibody to assess background. The images were captured at 60X magnification.

Western blot was performed using Anti-MCM7 Polyclonal Antibody (Product # PA5-79651) and a 80 kDa band corresponding to MCM7 was observed in all the tested cell models. Modified whole cell lysate (1% SDS) (30 µg lysate) of PC-3 (Lane 1), LNCaP (Lane 2), DU 145 (Lane 3), HeLa (Lane 4), MCF7 (Lane 5), MKN45 (Lane 6), SW480 (Lane 7), and Jurkat (Lane 8) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (0.5 µg/mL) and detected by chemiluminescence with Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4,000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).