Measuring peptide concentration
khatipovNO at NOuchicago.edu
Tue Mar 19 16:47:39 EST 2002
So these are cysteines that give some fluorescence to my non-aromatic
peptides! It's never too late to learn. Thanks for opening my eyes a little
bit wider, Artem :-)
"Artem Evdokimov" <AEVDOKIMOZ at cinci.rr.com> wrote in message
news:GBxl8.166129$Hu6.38723197 at typhoon.neo.rr.com...
> I disagree on one point : Expasy server does NOT calculate extinciton if
> don't have certain types of AA's in your sequence.
> For example 'AAAAAA' results in
> Extinction coefficients:
> As there are no Trp, Tyr or Cys in the region considered, your protein
> should not be visible by UV spectrophotometry.
> There's a reason for it - while some absorption at 280 will indeed be
> present, it won't be characteristic, and it would be overwhelmed by
> non-specific absorption of crud.
> Otherwise, way to go Emir :)
> "Emir Khatipov" <khatipovNO at NOuchicago.edu> wrote in message
> news:pyul8.311$s4.23478 at news.uchicago.edu...
> > Use Protparam tool at www.expasy.ch/tools to calculate molar extinction
> > coefficient of your peptide and use it for your spectrophotometric
> > measurements. Peptides absorb light at 280nm even if they don't contain
> > aromatic residues. I hope it is needless to say that you should use same
> > DMSO in PBS as a reference. If you believe there are impurities in your
> > samples, I would suggest determining absorption spectra of your peptide
> > solutions at ~200-350nm (or at least 250-330nm). If your sample is pure,
> > should see a nice peak at around 276-280nm. Be aware that peptide bond
> > absorbs at 214nm, but there might be a lot of interference from
> > you may have in your sample.
> > Emir
> > "JB" <j.bukczynskiDELETE_ME at utoronto.ca> wrote in message
> > news:b87a9ugslasbigp8lm4dhkqc4p0soe0se7 at 4ax.com...
> > > Hi,
> > >
> > > Does anyone know how to measure peptide concentration using a
> > > spectrophotometer? I have it dissolved in 10% DMSO in PBS. Any help
> > > would be appreciated. Thanks,
> > >
> > > Jacob
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