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I want to measure alkaline phosphatase activity using PNPP in my mutant bacteria strains, but all the protocols I found involve purification of the phosphatase (which I have no need of).

Does anyone know a good protocol which involves using only the supernatant, without purification of the enzyme? What is the preferred buffer for the reaction?

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I found this kit that appears to be compatible with biological samples. Rather than buying the kit you may also be able to imitate / modify their protocol to do the assay with your super natant.

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  • $\begingroup$ Thank you, but I don't think I can imitate the kit - they don't share the components of the buffers. It did give me some idea of how the protocol is done, so thank you. $\endgroup$ – Innab Dec 24 '11 at 7:46
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There's a paper from 1973 describing a very simple protocol:

Wolf PL, Von der Muehll E, Praisler K. 1973. A test for bacterial alkaline phosphatase: use in rapid identification of Serratia organisms. Clinical chemistry 19: 1248–9.

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