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
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.
LITAF Antibody dectects endogenous levels of total human LITAF protein.
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human LITAF protein. Antibodies were purified by peptide affinity chromatography.
LITAF (PIG7/SIMPLE) is protein that contributes to the regulation of the inflammatory cytokine TNF-α (1-3). LITAF was identified as a transcription factor involved in LPS-induced TNF-α expression through interaction with the TNF-α promoter (1,4). The same protein, called PIG7, was independently described in a model for p53 regulation (2). A putative alternative spiced form of LITAF, named SIMPLE, encodes a protein with a unique carboxyl terminus (3). Studies on LITAF-deficient mice demonstrate that LITAF plays a significant role in the regulation of several inflammatory cytokines in response to LPS (5). The regulation of LITAF can occur through phosphorylation by p38α via the TLR pathway that leads to its nuclear translocation (5). Mutation in the LITAF/SIMPLE gene has been associated with an autosomal dominant demyelinating form of Charcot-Marie-Tooth disease (6).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
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|2879S||100 µl (10 western blots)||$255.00.0|