From: John Philo <jphilo@mailway.com>
  To  : Joel Mackay <j.mackay@biochem.usyd.edu.au>
  Date: Fri, 5 Oct 2001 17:27:26 -0700

RE: light scattering

Joel,

These on-line light scattering detectors are quite, quite handy for
identifying the native state of oligomerization of proteins, the size of
aggregate species, and the stoichiometry of multi-protein complexes. Also
useful for characterizing degree of conjugation with things like PEG.

As you surmised, though, this is generally not a good approach to measuring
association constants, and generally would be used with purified proteins.
It is possible to get a true solution mass for an impure protein if it
elutes as a reasonably resolved peak on SEC---something that's pretty tough
to do any other way!

We have a number of example biotech/pharmaceutical applications on our web
site---see http://www.ap-lab.com/light_scattering.htm  You also might want
to see our review article in Analytical Biochemistry which has examples with
protein-protein complexes [Wen, J., Arakawa, T., and Philo, J. S. (1996).
Size-exclusion chromatography with on-line light-scattering, absorbance, and
refractive index detectors for studying proteins and their interactions.
Anal. Biochem. 240, 155-166.]

The technique is really pretty straightforward, and you've got the equipment
you need, so "just do it"!

John Philo
Alliance Protein Laboratories

-----Original Message-----
From: Joel Mackay [j.mackay@biochem.usyd.edu.au]">mailto:j.mackay@biochem.usyd.edu.au]
Sent: Friday, October 05, 2001 5:02 PM
To: rasmb@alpha.bbri.org
Subject: light scattering


hi everyone,
Does anyone have a feel for the utility of multi-angle light scattering for
the determination of molecular weights of proteins in solution? I have only
just come across it as two detectors plumbed into the end of an FPLC - a
light scatterer and an instrument for measuring refractive index. The
literature says that it can reveal MW without a shape dependence, although
it looks like it would be non-trivial if there was more than one species
present (that is, it doesn't look like one could use it easily for
determining association constants etc, just MW for a single purified
species). Seems pretty powerful to be able to get a feel for MW in-line for
each signal off your gel filtration run.
The instruments i saw were from Wyatt (miniDawn and Optilab DSP
refractometer).
cheers
joel

*****************************************************************
Dr Joel Mackay
ARC Research Fellow
Department of Biochemistry, G08
University of Sydney,
Sydney, NSW 2006
Australia
ph +61-2-9351-3906
fax +61-2-9351-4726
WWW: http://www.biochem.usyd.edu.au/staff/mackay/
Sydney Protein Group Website:
http://www.biochem.usyd.edu.au/spg/
****************************************************************