Hello All,
There is even more troubling thing with BSA. Just couple weeks
ago I tried to use BSA as a standard for MALDI of big proteins.
Out of curiosity I changed linear mode to reflector on my DE-STR.
You can imagine my suprice when I detected 2 almost baseline resolved
peaks - one with mass 66428 (apparently BSA) and another with
a mass 69848 (BSA precursor). And precursor peak was much
stronger than BSA peak. But in linear mode I saw only one relatively
simmetrical peak 66300. Speaking about purity of commercially
available proteins!
ma
> Hi Jim,
>
> We've had trouble getting the theoretical mass of BSA. From a
> thermobioanalysis application note, it's 66431 Da. However, if we go
> directly from the sequence in Swiss Prot, the mass is 66433 Da for the
> reduced form, and 66398 for the form with disulfides (17 of those). I
> guess we are dealing w/ the form with disulfides. Yet, if the
> accuaracy is 0.1% like Arnie said, it'd be hard to differentiate the
> two. We also found that if we used the BSA and RNase A to calibrate
> the double charge peak of BSA, it was off from calculated by either of
> the numbers above, which probably means the BSA we used has a
> different mass, as we are pretty confident about RNase A.
>
> Regards,
> Wei
>
>
> ----- Original Message -----
> From: "Jim Kerwin" <jlk39@cornell.edu>
> To: "Arnie Falck" <falck@pacbell.net>; <wuwei1@msu.edu>
> Sent: Wednesday, February 21, 2001 10:19 AM
> Subject: Re: Big Proteins on MALDI
>
>
> >
> > Good morning,
> >
> > Another option: Bovine albumin (66,431) and it's dimer (132,859).
> > If you can reliably get ESI spectra from proteins > 50 kDa, please
> > send the rest of us details. Regards,
> >
> > Jim Kerwin
> > Mass Spectrometry Facility
> > Cornell University
> >
> >
> > >Wei Wu,
> > >The best standard at fairly high mass that I am aware of is yeast
> > >enolase (Sigma sells it), but it's only about 46k. If you slightly
> > >overload the sample, you can see the proton-bound dimer at around
> > >92k, though.
> Accuracy
> > >somewhat depends on what instrument you are running, and more on
> > >how
> clean
> > >your protein is (narrow peaks=better mass accuracy) but I expect
> > ><0.1% on mine for this mass with a good sample. ESI will usually
> > >give you better
> mass
> > >accuracy at this mass, IF you can get the protein to run.
> > >Arnie Falick
> > >Applied Biosystems
> > >FalickAM@appliedbiosystems.com
> > >
> > >----- Original Message -----
> > >From: "Wei Wu" <wuwei1@msu.edu>
> > >To: "Recipients of ABRF List" <abrf@aecom.yu.edu>
> > >Sent: Monday, February 19, 2001 7:47 PM
> > >Subject: Big Proteins on MALDI
> > >
> > >
> > >> Hi Folks,
> > >>
> > >> I'm trying to do MALDI on a pretty big protein (90k). What are
> > >> the
> mass
> > >> standards for that range? Are there any matrixes other than
> > >> sinapinic
> acid
> > >> that are worth trying for this big protein? Or there are some
> > >> special
> > >sample
> > >> preparation methods? Is it true that I should not expect a good
> > >> mass
> > >accuracy
> > >> at this high end?
> > >>
> > >> Thanks in advacne!
> > >>
> > >> Wei Wu
> > >>
> > >> --
> > >> 401 BCH
> > >> MSU
> > >> E. Lansing, MI 48824
> > >> 1-517-353-6767
> > >>
> > >>
> >
> >
>
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