[RASMB] AUC measurements in CsCl?

[Borries Demeler]

On Fri, Jun 7, 2019 at 4:19 AM Holger Strauss <HolgerMStrauss at gmx.de> wrote:

>
> When using CsCl2: thorough, thorough dialysis, vbar measurements and very
> accurate concentration measurements.
>
> Just something to be aware of here: Any vbar measurement not done in an
appropriate density matching experiment will not reveal the central point,
i.e, the vbar heterogeneity expected from the mixture of labeled and
unlabeled molecules. For example, a densitometer will give you a
weight-average which would be wrong for either species. In a sense, the
CsCl buoyant density equilibrium gradient experiment is almost perfect to
reveal this difference. I say almost, because you would measure a vbar that
is related to the hydration seen in a local CsCl concentration, which
differs for each DNA and may be different because of this (presumably less
hydration and more Cs bound in the more dense species, which will settle in
the higher density and higher concentration CsCl environment). And of
course, this vbar would also not be the same you would see under
physiological conditions. For highly charged polyanions like DNA the vbar
would be super sensitive to ionic strength. Since this vbar difference is
expected to lead to different peaks in the density gradient, this would be
evidence that the difference is big enough to be relevant.

-b.
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