ATP EXTRACTION; ATPASE INHIBITORS

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Martin - 06 May 2004 12:03 GMT

Hey,

I will be really gratefull if somebody tell how extract ATP from human
cells.
There is also another problem with ATPases - so after extraction we
should use ATPases inhibitors.
Maybe somebody knows which inhibitors are posible to use?
BEST WISHES!

Marcin

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Dr Engelbert Buxbaum - 10 May 2004 06:23 GMT

> Hey,
>
[quoted text clipped - 4 lines]
> Maybe somebody knows which inhibitors are posible to use?
> BEST WISHES!

ATPases first: Plasma membrane Na/K-ATPase is inhibited specifically
with ouabain (0.1 mM), F-type ATPases with NaN3 (3 mM). Depending on
what you want to do, vanadate can inhibit quite a few ATPases, but is
difficult to get rid of later. You may also wish to work at a slightly
acidic pH.

Isolation method depends on what you want to do: preparative isolation
of ATP (forget it: yeast is a much cheaper source) or determination of
ATP-concentration, e.g. with luciferase. In the latter case, no
isolation is actually required, a simple cell lysate will do.

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Martin - 10 May 2004 15:23 GMT

Thanks a lot! I am doing according to what you have proposed( simple
cell lysate and it works ,I study ATP amount in cells with
luciferin-luciferase assay),
kindly regards!
Marcin

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Natural Science Forum / Biology / Biology / May 2004

ATP EXTRACTION; ATPASE INHIBITORS