pertusis toxin - making a uniform suspension

John Hines hines at pharm.med.upenn.edu
Wed Aug 8 09:32:21 EST 2001


I want to use pertusis toxin to inactivate Gi family members in transfected
COS cells to investigate some signaling pathways.  I ordered the pertusis
toxin from Calbiochem (cat. no. 516560), and it comes as a lyophilized
solid to be reconstituted in water.  Both the label on the vial AND the
catalog description mention that I should obtain a "uniform suspension"
before using the toxin on cells.  I figured that should be easy enough.

As it turns out (to my surprise), the toxin forms a cloudy, particulate
suspension when water is added to the lyophilized powder.  Usually
lyophilized proteins go into solution very quickly and easily, but not so
here.  I tried vortexing the hell out of it, but the clumps of toxin
remain.

I don't know if the toxin would stand up to sonication; and I don't want to
add a detergent, since that will also get applied to the cells when I add
the toxin to them.


Does anyone who routinely works with pertusis toxin have any suggestions on
how to make a "uniform suspension"?


John

hines at pharm.med.upenn.edu




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