Hi,
I am interested in using the thermofluor to assess the stability of my protein
in different buffers. Can anyone recommend a vendor that supplies buffer
screens, possibly in 96 well format?
Not crystallization buffers, just ordinary storage buffers.
Thanks
brenda
Quoting Andreas Förster <[EMAIL PROTECTED]>:
Dear Kornelius,
I found the idea of doing Thermofluor on membrane proteins really
intriguing - for identifying the best buffer and detergent, secondary
detergents, even for checking crystallization drops that stayed
clear. (This latter experiment should theoretically be possible with
large drops, even though you'd be working very near the limits
claimed in the publications.)
In the end, I didn't find the method too useful because of noise
issues due to detergent and exposed hydrophobic portions of the
protein. I always felt that, in order to get reasonable signal, I'd
have to use protein at unreasonable concentrations.
The method works much better for soluble protein, though I can't tell
you a success story where it led to crystallization that seemed
impossible before.
Andreas
Kornelius Zeth wrote:
Dear all,
a question very related to the discussion before. I have been
reading the papers about the thermofluor experiment with great
interest. I wonder what people think about the underlying
principles/ideas and the success that the method yielded in their
own labs for crystallization or related purposes?
Has anybody used this method with membrane proteins in order to find
out the stability of the protein in the presence of a second
detergent?
Is the method limited to this certain dye (sypro orange)?
Have a nice day
Kornelius
P.S.: I will make a summary of all opinions.
----------------------------------------------
Kornelius Zeth
Max Planck Institute for Developmental Biology
Dept. Protein Evolution
Spemannstr. 35
72076 Tuebingen, Germany
[EMAIL PROTECTED]
Tel -49 7071 601 323
Fax -49 7071 601 349
