A simple cell-alignment protocol for sedimentation velocity analytical ultracentrifugation to complement mechanical and optical alignment procedures

European Biophysics Journal : EBJ
Guy ChannellS E Harding

Abstract

In establishing the sources of data variability within sedimentation velocity analysis in the analytical ultracentrifuge and their relative importance, recent studies have demonstrated that alignment of the sample cells to the centre of rotation is the most significant contributing factor to overall variability, particularly for the characterisation of low levels of protein aggregation. Accurate mechanical and optical alignment tools have been recently designed. In this study, we (1) confirm the effect of misalignment observed by others on the estimated amounts of bovine serum albumin (BSA) monomer and dimer, and the sedimentation coefficient value for the BSA dimer; and (2) demonstrate the high performance of a mechanical alignment tool and the usefulness of a simple and complementary enhanced manual alignment protocol which should be useful for situations where these tools are not available.

References

Dec 7, 2007·Biotechnology & Genetic Engineering Reviews·Ulf NobbmannStephen E Harding
Dec 29, 2010·Methods : a Companion to Methods in Enzymology·John P Gabrielson, Kelly K Arthur
Sep 5, 2012·Carbohydrate Polymers·Ali Saber AbdelhameedStephen E Harding
Feb 18, 2014·Analytical Biochemistry·Huaying ZhaoPeter Schuck

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Citations

Sep 16, 2018·European Biophysics Journal : EBJ·Olwyn ByronTrushar R Patel
Jan 6, 2021·European Biophysics Journal : EBJ·Huaying ZhaoPeter Schuck
Nov 14, 2020·European Biophysics Journal : EBJ·Vlad DinuIan D Fisk
Jun 8, 2021·Molecular Therapy. Methods & Clinical Development·Harshit KhasaChunlei Wang

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Methods Mentioned

BETA
dynamic light scattering

Software Mentioned

Nanolytics
SEDFIT

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