[RASMB] s value and classical sucrose gradient separation

[david.hayes at boehringer-ingelheim.com]

Hi John,

I never got round to either before I left MedImmune, but I was convinced back then that elutriation or A4F would work better and more robustly for cells and nano-particles than density gradients ever could.

In elutriation sedimentation is countered by fluid flow in a special centrifuge cell.  I believe that the exact speed is chosen empirically.

The new A4F setup with an electric field could also result in a more pure sample since junk with the same hydrodynamic radius  but a different charge could be separated from the peak of interest by the electric field.

Beckman and Wyatt, respectively, can give protocols for these procedures.

Kind Regards,

David


From: RASMB [mailto:rasmb-bounces at list.rasmb.org] On Behalf Of John Sumida
Sent: Thursday, November 29, 2018 3:36 PM
To: RASMB
Subject: [RASMB] s value and classical sucrose gradient separation

Dear RASMB,

How would one use the sedimentation value to design a classical sucrose gradient experiment to extract and then purify a species of interest?

This question is in relation to the characterization and isolation of nanoparticles.

I imagine it would be using the observed sedimentation value obtained experimentally and calculating r to determine the radius at a given buoyance ratio as per

[cid:image001.png at 01D487FB.D78892E0]
Hirst_Cox_Biochem_J_1976_159 (see attached).

However, if there is a standard protocol in current use, I would be interested in a reference if appropriate.

Thank you for considering my question.

Best regards,
John Sumida
Molecular Analysis Facility
University of Washington



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