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washing of phosphorylated plasma protein

washing of phosphorylated plasma protein - Basic Lab Protocols and Techniques

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Old 09-18-2007, 09:18 AM
Pipette Filler
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Default washing of phosphorylated plasma protein



All proteins in a plasma sample is precipitated with 35% ACETONE/WATER and centrifuged. Then I want to wash the protein pellet one or several times to make sure that there is no free phosphate (from the plasma) associated with the pellet.
After this I dissolve the pellet in 1 M NaOH and heat to 70C to release all (alkali labile) protein bound phosphate for quantification. My question:
What buffer should I wash my protein pellet with that ensures complete removal of free phosphate and plasma but at the same time without releasing serine-bound phosphate?
Many thanks to you who can help a PhD student in analytical chemistry with little experience of working with blood and proteins.
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