2 replies to this topic

[asias]

Hi everyone

I work with phosphor-specific antibodies as a control a use alkaline phosphatase. Phosphatase work well when I use them directly on the blot or cell lisate but do not work when I use then in IP. The mine different between IP and the cell lisates is that IP is washed with PBS and as a results complex protein-antibodies-proteinA is in PBS.

Do you know that PBS can inhibit alkaline phosphatise? It is possible?

Edited by asias, 15 January 2010 - 04:01 AM.

[mdfenko]

yes. free phosphate will inhibit ap.

[bob1]

Use a tris- buffered wash instead of PBS.