[RASMB] DTT for equilibrium runs?

[Borries Demeler]

Hi Debby,

DTT as well as other reductants absorb in the UV. In addition, reductants
tend to change their extinction coefficient between reduced and oxidized
states. Take a look at this reductant table:

http://www.cauma.uthscsa.edu/ (click on "Resources", then "Reductants")

The more your buffer absorbs, the less dynamic range in the absorbance
is available to your protein. The XLA's spectrophotometer is only good
between about 0-1 OD, so if you use up half of this for your buffer
absorbance (essentially, raise the baseline absorbance) you don't have
much left for sample absorbance.

There are some alternatives: If you have to have reductant present
you can measure pretty much without problem at 280 nm if you replace
DTT with TCEP.

Good luck,
-Borries


> 
> Hi, everyone,
> 
> I just heard recently that it's not good to have DTT in your buffer for 
> doing equilibrium runs.  Can someone explain to me why that would be?  Or 
> share stories of DTT use?
> 
> Thanks very much,
> 
> Debby
> 
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