Western blot analysis of extracts from various cell lines using AHSA1 Antibody.
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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
AHSA1 Antibody recognizes endogenous levels of total AHSA1 protein.Species Reactivity:
Human, Mouse, Rat, Monkey
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu242 of human AHSA1 protein. Antibodies are purified by protein A and peptide affinity chromatography.
Activator of 90 kDa heat shock protein ATPase homolog 1 (AHSA1 or AHA1) is a 42 kDa HSP90 co-chaperone that stimulates intrinsic HSP90 ATPase activity in eukaryotic cells to facilitate client protein folding and activation (1,2). Initially, HSP90 forms a ternary complex with a nascent client protein such as v-Src, c-Raf or CFTR, and the co-chaperone protein CDC37 (3,4). HSP90 then cycles through a series of conformational changes facilitated by ATP that results in the release of CDC37 and recruitment of AHSA1 - a process that is dependent on tyrosine phosphorylation of HSP90 and CDC37 (5). More recently, AHSA1 has been shown to have unique interactions with additional client proteins involved in RNA splicing and DNA repair, suggesting that AHSA1 may have also have a scaffolding role in recruiting client proteins to HSP90 (6).
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