[RASMB] Optics check

Thu Apr 5 12:05:12 PDT 2007

Hey Borries
I did learn this test from you! It has never been this way. I try my best to
take care of the machine but it is a common instrument and well...I can't be
the AUC police here....these problems started when someone else was using it
and well...it's been downhill ever since.
I tried Dr. Correia's intensity versus radial position test. The intensity is ~
5200 at 6.0 cm and tapers off to ~ 4600 at 7.0 cm at 280 nm.
The monochromator is my suspect too. Of course, I call the Beckman Service
people and they sneak in, change something (the lamp power source, the slit
assay), mention that they did some autoscan test etc., and vanish before I even
know they were here.
BTW, any Beckman Representative on this forum?!! Ooops....
I'll be glad to hear other views on this problem.
Mitra

Quoting Borries Demeler <demeler at biochem.uthscsa.edu>:

> Mitra,
>
> This is the check I recommend you run to make sure your optics are
> properly aligned and tuned. Your system doesn't seem to be.
>
> If this results is obtained after cleaning the lamp and windows I would
> suspect a problem with your monochromator. I think it is time to call
> service. What you describe does not sound like a well-functioning machine.
>
> -Borries
>
> >
> > That was a prompt reply.
> > I did the intensity scans on a windowless cell. Basically air vs. air. I
> haven't
> > tried intensity scans on velocity mode at various wavelengths yet. In fact
> I am
> > heading down to do this right now.
> > Mitra
> >
> > Quoting John Correia <jcorreia at biochem.umsmed.edu>:
> >
> > > Sounds like you are doing intensity scans of real samples?  1st try
> intensity
> > > scans in velocity mode on an empty cell at low speed (3-4K) or on an
> empty
> > > hole, air vs air, where you scan the whole cell from 5.8 to 7.2 cm.  Vary
> the
> > > wavelength for each scan.  Machine spec is 20% variation in linearity &
> is
> > > done at 280 nm.   Often the intensity drops off near the top ~ 5.85 cm
> which
> > > is a function of the alignment of the flash lamp - slight rotation can
> make
> > > it better although not by much with the new lamp design.  This is your
> > > baseline for intensity.  With a real sample vs buffer the sample will
> absorb
> > > more light & of course they will not overlap.  The log ratio is the
> > > absorbance of the sample & in your case is must be > 1.5 - 2.0 OD at that
> > > wavelength?
> > >
> > >
> > > -------------------------------------------------------------------
> > >  Dr. John J. "Jack" Correia
> > >  Department of Biochemistry
> > >  University of Mississippi Medical Center
> > >  2500 North State Street
> > >  Jackson, MS  39216
> > >  (601) 984-1522
> > >  fax (601) 984-1501
> > >  email address: jcorreia at biochem.umsmed.edu
> > >  homepage location: http://biochemistry.umc.edu/correia.html
> > >  dept homepage location:    http://biochemistry.umc.edu/
> > > -------------------------------------------------------------------
> > >
> > >
> > >
> > >
> > > >>> <mitrana at mail.utexas.edu> 04/05/07 11:12 AM >>>
> > >
> > > Hi all
> > > I did a intensity versus wavelength scan at three radial positions - 5.8,
> > > 6.5,
> > > and 7.2 cm. At 6.5 cm, it was as I expected with a 230 nm peak (I ~ 10000
> > > units) and half that at 527 nm. However, at 5.8 cm, the 230 peak was
> almost
> > > zilch and only the 527 prominent. And at 7.2 cm, 230 peak had an
> intensity of
> > > ~
> > > 13000 units and 527 half that. However, the sample and reference did not
> > > overlay. The sample intensity was virtually zero.
> > >
> > > I am a little lost right now as to what is going on. Are there anymore
> tests
> > > I
> > > can run to identify the exact problem?
> > >
> > > Mitra
> > >
> > >
> > > --
> > > Mitra S. Rana
> > > Graduate Student
> > > Institute for Cellular and Molecular Biology
> > > 2500 Speedway, UT-Austin
> > > Austin, TX-78712
> > > _______________________________________________
> > > RASMB mailing list
> > > RASMB at rasmb.bbri.org
> > > http://rasmb.bbri.org/mailman/listinfo/rasmb
> > >
> > >
> > >
> >
> >
> > --
> > Mitra S. Rana
> > Graduate Student
> > Institute for Cellular and Molecular Biology
> > 2500 Speedway, UT-Austin
> > Austin, TX-78712
> > _______________________________________________
> > RASMB mailing list
> > RASMB at rasmb.bbri.org
> > http://rasmb.bbri.org/mailman/listinfo/rasmb
> >
>

-- 
Mitra S. Rana
Graduate Student
Institute for Cellular and Molecular Biology
2500 Speedway, UT-Austin
Austin, TX-78712