Western blot analysis of extracts from various cell lines using MTMR14 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
MTMR14 Antibody recognizes endogenous levels of total MTMR14 protein.Species Reactivity:
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Arg500 of human MTMR14 protein. Antibodies are purified by protein A and peptide affinity chromatography.
Myotubularin-related protein 14 (MTMR14), also known as Jumpy, is a myotubularin-related phosphoinositol-3-phosphate (PI3P) phosphatase (1). Mutations in the MTMR14 gene have been associated with centronuclear myopathy (1). MTMR14 deficiency in mice leads to altered calcium homeostasis and muscle disorders (2). MTMR14 has also been shown to play a role in autophagy, a process that is highly regulated by phosphatidylinositides through the type III PI3K, Vps34 (3). MTMR14 was localized to autophagic isolation membranes and early autophagosomes (3). In these studies, MTMR14 inhibited autophagy and mutations of MTMR14 associated with centronuclear myopathy were also defective in autophagy inhibition. In zebrafish, MTMR14 knockdown was shown to increase the number of autophagosomes, suggesting that its activity is associated with an inhibition of autophagy (4).
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