For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
Monoclonal antibody is produced by immunizing animals with recombinant protein specific to the EDB domain of human Fibronectin/FN1 protein.
Fibronectin is a multi-domain extracellular matrix (ECM) protein. The protein uses its different domains to bind distinct ECM components such as collagens, growth factors, and cell surface integrins to carry on its functions (1). Fibronectin has been implicated in many essential biological processes, including tissue repair, fibrosis, and tumor development (1,2). There are many fibronectin isoforms. Plasma fibronectin is synthesized by hepatocytes and exists as a compact, inactive conformation in the bloodstream. It is a major component of fibrin clots. Upon binding to integrins or other cell surface receptors, fibronectin switches to an extended conformation, and exposes its function domains to activated extracellular matrix assembly (3). Fibroblasts, endothelial cells, and many types of cancer cells have also been shown to synthesize cellular fibronectin isoforms. Among them, the EDA or EDB isoforms are particularly highly expressed during fibrosis and cancer development, and are potential diagnostic and therapeutic targets (4-6).
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