SEAP assay

[Mike Holloway]

>In article <thompsop-2905961612510001 at 203.0.141.239>,
>thompsop at burnet.mbcmr.unimelb.edu.au (Phillip R Thompson) wrote:
>
>> I am using Clontech's Great Escape SEAP Genetic reporter system & am
>> having a major problem with reproducibility.  Some of my duplicates differ
>> by as much as 300%!  The assays are being carried out essentially as
>> described in the manual & read in 96-well black plates in a Wallach plate
>> reader.  I'm being very careful with pipetting & I don't believe pipetting
>> error could result in such large errors anyway.  Any help would be
>> appreciated!

Assuming that you've deactivated the other phosphatases (65 to 70
degrees for at least one hour, more's OK) you need to normalize with a
cotransfected reporter.  The assay's very sensitive.  Small
differences will definitely produce variability.

Mike Holloway
holloway at cris.com