TCA precipitation of proteins

Deef Suus deefsuus at casema.net
Mon Jul 17 15:10:16 EST 2000


Than what is the best way to concentrate my samples?
I purified proteins using affinity chromatography. After the column step I
have my desired transporter protein.
I have about 30ml of samples with a protein concentration as low as 10ug/ml.
There are also lots of salts in there (eg. DTT, 3.5M NaCl, lactic acid,
n-octylglucoside). I want to determine the protein concentration. Right now
I am using an BCA assay. But the problem is that DTT (and NaCl) interferes.
I also want to build them into liposomes to do some transport studies. Right
now I am using so-called concentrators. But the problem is with this amount
of sample you are centrifuging the whole day (which is not very prefereable
for the proteins; but I guess the TCA is even worse.. :)  )

Does anybody have any idea.

"Chris LaRosa" <clarosa at biocomp.unl.edu> wrote in message
news:397321E9.EE62DCE5 at biocomp.unl.edu...
>
>
> Suzanne Hermeling wrote:
> >
> > Hi
> >
> > I have a very brief question:
> > When you perform a TCA precipitation of proteinsamples, do the proteins
> > denature?
> > Or can you "rebuild" them in a phospholipid membrane again?
> >
> > Suzanne
>
> They are definitely denatured!... Thats why TCA ppt is recommended for
> protease inactivation.







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