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Technical Data Sheet

EL7331

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Human Endothelial Calyculin A Control
Lysate
Price
Size

$75
100 μl

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Application
Dilution


WB
20 μl/lane



End user should determine optimal dilution for their particular applications and experiments.Western blot membranes were incubated with diluted antibody in 5% non-fat milk, PBS, 0.04% Tween20 for 1hour at room temperature.
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Background
Calyculin A is a serine/threonine phosphatase inhibitor that inhibits the activity of protein phosphatases PP1 and PP2A. Cells treated with calyculin A for 30 minutes can undergo significant threonine phosphorylation compared to control cells, as shown by western blotting using anti-Phosphoserine/threonine, cat.# PP2551.

Confluent cultures of human umbilical cord endothelial cells were serum starved 2 hrs at 37°C. Cells were then either left untreated (Cat.# EL7331) or treated with Calyculin A (100 nM) for 30 minutes at 37°C (Cat # EL7341). Cells were lysed in 1% SDS, 1 mM sodium ortho-vanadate, 1 mM sodium fluoride, 10 mM Tris (pH 7.4) buffer. Protein concentration was determined using the BCA method (Pierce) before diluting to final concentration and buffer.

Buffer and Storage
Cell Lysates are supplied at a concentration of 1 mg/ml in electrophoresis sample buffer (62.5 mM Tris pH 6.8, 2% SDS, 5% glycerol, 0.003% bromophenol blue, 0.9% β-mercaptoethanol). Store at –20°C. Do not boil or dilute. Stable for 1 year.
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