Western blot was performed using anti-Flotillin 2 Polyclonal Antibody (Product # PA5-21296) and a 53 kDa band corresponding to FLOT2 was observed in HeLa and A-431 but not in Molt-4 which is reported to be negative. Whole cell extracts (30 µg lysate) of HeLa (Lane 1), A-431 (Lane 2) and Molt-4 (Lane 3) were electrophoresed using NuPAGE™ 10% Bis-Tris Protein Gel (Product # NP0301BOX). 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 (1:1000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).