Immunocytochemical analysis of ARF6 in HepG2 cells using a ARF6 Monoclonal antibody (Product # MA5-32558) as seen in red. The nuclear counter stain is DAPI (blue). Cells were fixed in paraformaldehyde, permeabilised with 0.25% Triton X100/PBS.

Western blot was performed using Anti-ARF6 Recombinant Rabbit Monoclonal Antibody (JF101-2)(Product # MA5-32558) and a 20kDa band corresponding to ARF6 was observed across tissues (Fig.a) and cell lines (Fig.b) tested except in Mouse Skeletal Muscle and Bone Marrow and MCF 10A which are reported to be either negative or low in expression for ARF6. Uncharacterized bands (*) were also observed in cell lines and tissues tested. Tissue extracts (30 µg lysate) of Mouse Lung (Lane 1), Mouse Kidney (Lane 2), Mouse Liver (Lane 3), Mouse Skeletal Muscle (Lane 4) and Mouse Bone Marrow (Lane 5) (Fig.a) and Whole cell extracts (40 µg lysate) of MCF 10A (Lane 1), MCF7 ( Lane 2) and HEK-293 (Lane 3) ( Fig.b) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). 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 (1:2000 dilution for tissues, 1:3000 for cell lines) 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).