Western Blot was performed using Anti-CD133 Monoclonal Antibody (5E3) (Product # MA5-18323) and a 133 kDa band corresponding to Prominin-1 was observed across cell lines tested except HeLa and SK-O-V3 which are reported to be low or negative for CD133 expression. Membrane enriched extracts (30 µg lysate) of NTERA-2 cl.D1 (Lane 1), Caco-2 (Lane 2), HT-29 (Lane 3), HeLa (Lane 4) and SK-O-V3 (Lane 5) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23002) 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-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).