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.
HTATIP2/TIP30 Antibody recognizes endogenous levels of total HTATIP2 protein.
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding His231 of human HTATIP2 protein. Antibodies are purified by protein A and peptide affinity chromatography.
The HIV-1 Tat interactive protein 2 (HTATIP2, TIP30, CC3) is an oxidoreductase that was originally identified as a metastatic tumor suppressor and Tat-mediated proapoptotic gene transcription cofactor (1,2). HTATIP2 protein contains a short-chain dehydrogenase (SDR) domain and a NADPH binding motif important for HTATIP2 interaction with importins and inhibition of nucleocytoplasmic transport (3,4). Research studies demonstrate that induced overexpression of HTATIP2 predisposes cells to apoptosis by inhibiting the nuclear transport of important signaling proteins (e.g. p53, activated notch1) and several key targets of the DNA repair process (5-7). HTATIP2 is part of a protein complex, with Rab5a, endophilin B1, and ACSL4, that may regulate EGFR receptor endosomal trafficking, degradation, and cytoplasmic/nuclear signaling (8,9). Silencing of HTATIP2 promotes tumor cell survival under low glucose conditions by inducing increased expression of mitochondrial respiratory proteins and glucose metabolic enzymes (10).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. Tween is a registered trademark of ICI Americas, Inc.
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|14614S||100 µl (10 western blots)||$ 255.0|