Immunofluorescent analysis of Villin in methanol-fixed HeLa cells using a Villin polyclonal antibody (Product # PA5-22072) (Green) at a 1:500 dilution. Alpha-tubulin filaments were labeled with Product # PA5-29281 (Red) at a 1:2000.

Western blot was performed using Anti-Villin Polyclonal Antibody (Product # PA5-22072) and a 92kDa band corresponding to Villin was observed along with uncharacterized bands (*) across cell lines and tissues tested. Whole cell extracts (30 µg lysate) of Caco-2 (Lane 1), COLO 205 (Lane 2), Jurkat (Lane 3) and PANC-1 (Lane 4) as seen in Fig (a).Tissue extracts of Mouse Liver (Lane 1), Mouse Colon (Lane 2) and Mouse Jejunum (Lane 3) as seen in Fig (b) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Relative expression of Villin was observed high in Caco-2 and COLO 205 and low to negative in Jurkat and PANC-1 as seen in Fig (a), similarly expression of Villin was high in Mouse Jejunum and low to negative in Mouse Colon and Mouse Liver as seen in Fig (b). 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 (Heavy Chain) 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).