From: Lisa Joss <ljoss@hci.utah.edu>
  To  : David Scott <djs17@york.ac.uk>
  Date: Mon, 25 Jan 1999 12:19:39 -0700

Re: absorbance values

Hi Dave,

I'm inclined to think that the steepness of the concentration gradient
is more of a deciding factor than the absolute OD. In the past I have
found that steep concentration gradients, like those sometimes seen in
equilibrium runs, can result in a redirection of  light and thus anomalous
absorbance readings toward the base of the channel.  This effect is a
consequence of Weiner skewing. The problem can be quite profound and
yet still give data that at first inspection look quite reasonable. In
light of this, I always include a scan at a non absorbing wavelength
(usually at about 360nm for most proteins). This scan should be flat over
the radius for which there is no light redistribution (assuming there
are no finger prints or scratches on the window). Often times this scan
will have a significant wave of positive and negative absorbance toward
the base. I then make sure that I am not using any data in my 280nm scan
that includes this region of "up-down" stuff at the end of the 360nm.

I didn't really answer your question but I hope this is helpful to
you anyway.

Cheers
Lisa

David Scott wrote:

> Hia RASMBers
>
> What is the highest practical OD reading that I can use in my sedimentation
> equilibrium scans before it becomes non-linear. I've been quoted values ranging
> from 1.0 to 2.5 ODs.
>
> Can anyone clear this up?
>
> Dave Scott.
>
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> * Dr. David Scott               *
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> * University of York            *
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--
______________________________________________________________________________

Lisa Joss, Ph.D.
Huntsman Cancer Institute                  voice: (801) 585-5359
University of Utah                         fax:   (801) 585-3833
50 N. Medical Drive #4A437                  email: Ljoss@hci.utah.edu
Salt Lake City, UT, 84132
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