Western blot analysis of extracts from SF-539 and ACHN cells using COL5A1 Antibody (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). As expected, ACHN cells are negative or low for COL5A1 expression.
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
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
COL5A1 Antibody recognizes endogenous levels of total COL5A1 protein.
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro364 of human COL5A1 protein. Antibodies are purified by peptide affinity chromatography.
The Extracellular Matrix (ECM) is a complex network of macromolecules that provides structural tissue support to cells in the basement membrane and interstitial matrix. It is composed of many molecules including proteins, glycoproteins, proteoglycans, and polysaccharides (1,2). One of the major proteins that comprises the ECM, and the human body, is collagen. Collagens are a large family of proteins. They are trimeric molecules comprised of three alpha polypeptide chains that form a triple helix structure that is characteristic of all collagens (3). The large family of collagens is divided into three sub groups: the fibrillar collagens, the non-fibril forming collagens, and the fibril-associated collagens. These sub groups differ in their structure and supramolecular assembly (3).
COL1A5 is a minor fibrillar collagen expressed broadly across tissues. It participates in bone and cartilage development, and defects in the COL1A5 gene leads to Ehlers-Danlos Syndrome, a condition that affects the integrity of bones, cartilage and skin (4,5). Several proteomics, gene profiling, and biochemical studies have indicated that aberrant COL5A1 may play a malignant role in various types of cancer including breast, renal, colon, prostate, lung, and others (6-11).
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