[RASMB] AUC measurements in CsCl?

[Laue, Thomas]

Bo is correct. There are a great set of papers on using CsCl gradients by Vinograd.
I would just add that vbar also depends on the salt type (particularly cation type).
Best wishes,
Tom

From: RASMB <rasmb-bounces at list.rasmb.org> On Behalf Of Borries Demeler
Sent: Friday, June 7, 2019 8:55 AM
To: Holger Strauss <HolgerMStrauss at gmx.de>
Cc: RASMB <rasmb at list.rasmb.org>; Mats Leeman <mats at solveresearch.com>
Subject: Re: [RASMB] AUC measurements in CsCl?

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