Western blot analysis of extracts from various cell lines using COL4A1 Antibody (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). The absence of COL4A1 expression in HCT 116 cell extracts is consistent with molecular and proteomic expression profiling data, confirming specificity of the antibody.
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 datasheet or 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 November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised October 2016
Protocol Id: 10
COL4A1 Antibody recognizes endogenous levels of total COL4A1 protein.Species Reactivity:
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Asp536 of human COL4A1 protein. Antibodies are purified by peptide affinity chromatography.
Type IV collagens support extracellular matrix (ECM) function by forming a cross-linking network in the basement membrane zone. There are six different type IV collagens that are encoded by the following genes: COL4A1, COL4A2, COL4A3, COL4A4, COL4A5, and COL4A6. Among them, COL4A1 and COL4A2 are the most common collagens found in the basement membrane, and are usually used as a marker of the basement membrane dynamic stability (1). The type IV collagen fiber exists as a trimeric complex comprised of two molecules of COL4A1 and one molecule of COL4A2. The N-terminal 7S domains of the protein function on intermolecular cross-linking and C-terminal non-collagenous domains (NCI) are responsible for heterotrimer assembly. The trimeric fiber further associates to form cross-linked sheet structure by NCI/NCI and 7S/7S interaction (2). COL4A1 plays a critical role in maintaining basement membrane integrity for normal tissue functions. Genetic mutations of COL4A1 and COL4A2 have been linked to cardiovascular disease, cerebral small vessel disease, and glomerulocystic kidney disease (3-5).
Explore pathways + proteins related to this product.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Tween is a registered trademark of ICI Americas, Inc.