Western blot analysis of extracts from various cell types using Chronophin/PDXP (C85E3) Rabbit mAb.
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Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. 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
Chronophin/PDXP (C85E3) Rabbit mAb detects endogenous levels of total chronophin/PDXP protein.Species Reactivity:
Human, Mouse, Rat, Hamster, Monkey, Bovine
Monoclonal antibody is produced by immunizing animals with recombinant mouse MBP-chronophin.
Cofilin is a conserved actin-severing protein required for processes that rely on actin dynamics, including cytokinesis and cell motility (reviewed in 1). Regulation of actin dynamics requires the controlled cycling between the phosphorylated and unphosphorylated forms of cofilin (2). The severing activity of cofilin is inhibited by LIMK or TESK phosphorylation at the conserved amino-terminal Ser3 of cofilin (3,4). Slingshot (SSH) phosphatase, for which there have been three mammalian isoforms identified, dephosphorylates cofilin in vivo (5). Chronophin (CIN, PDXP) is a haloacid dehalogenase phosphatase that also dephosphorylates cofilin. Alteration of CIN activity through overexpression of either the wildtype or phosphatase-inactive mutant CIN interferes with actin dynamics, cell morphology and cytokinesis (6).
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U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.