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A Multilaboratory Comparison of Calibration Accuracy and the Performance of External References in Analytical Ultracentrifugation

Lookup NU author(s): Dr Alexandra Solovyova

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This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


Abstract

Analytical ultracentrifugation (AUC) is a first principles based method to determine absolute sedimentation coefficients and buoyant molar masses of macromolecules and their complexes, reporting on their size and shape in free solution. The purpose of this multi-laboratory study was to establish the precision and accuracy of basic data dimensions in AUC and validate previously proposed calibration techniques. Three kits of AUC cell assemblies containing radial and temperature calibration tools and a bovine serum albumin (BSA) reference sample were shared among 67 laboratories, generating 129 comprehensive data sets. These allowed for an assessment of many parameters of instrument performance, including accuracy of the reported scan time after the start of centrifugation, the accuracy of the temperature calibration, and the accuracy of the radial magnification. The range of sedimentation coefficients obtained for BSA monomer in different instruments and using different optical systems was from 3.655 S to 4.949 S, with a mean and standard deviation of (4.304 +/- 0.188) S (4.4%). After the combined application of correction factors derived from the external calibration references for elapsed time, scan velocity, temperature, and radial magnification, the range of s-values was reduced 7-fold with a mean of 4.325 S and a 6-fold reduced standard deviation of +/- 0.030 S (0.7%). In addition, the large data set provided an opportunity to determine the instrument-to-instrument variation of the absolute radial positions reported in the scan files, the precision of photometric or refractometric signal magnitudes, and the precision of the calculated apparent molar mass of BSA monomer and the fraction of BSA dimers. These results highlight the necessity and effectiveness of independent calibration of basic AUC data dimensions for reliable quantitative studies.


Publication metadata

Author(s): Zhao HY, Ghirlando R, Alfonso C, Arisaka F, Attali I, Bain DL, Bakhtina MM, Becker DF, Bedwell GJ, Bekdemir A, Besong TMD, Birck C, Brautigam CA, Brennerman W, Byron O, Bzowska A, Chaires JB, Chaton CT, Colfen H, Connaghan KD, Crowley KA, Curth U, Daviter T, Dean WL, Diez AI, Ebel C, Eckert DM, Eisele LE, Eisenstein E, England P, Escalante C, Fagan JA, Fairman R, Finn RM, Fischle W, de la Torre JG, Gor J, Gustafsson H, Hall D, Harding SE, Cifre JGH, Herr AB, Howell EE, Isaac RS, Jao SC, Jose D, Kim SJ, Kokona B, Kornblatt JA, Kosek D, Krayukhina E, Krzizike D, Kusznir EA, Kwon H, Larson A, Laue TM, Le Roy A, Leech AP, Lilie H, Luger K, Luque-Ortega JR, Ma J, May CA, Maynard EL, Modrak-Wojcik A, Mok YF, Mucke N, Nagel-Steger L, Narlikar GJ, Noda M, Nourse A, Obsil T, Park CK, Park JK, Pawelek PD, Perdue EE, Perkins SJ, Perugini MA, Peterson CL, Peverelli MG, Piszczek G, Prag G, Prevelige PE, Raynal BDE, Rezabkova L, Richter K, Ringel AE, Rosenberg R, Rowe AJ, Rufer AC, Scott DJ, Seravalli JG, Solovyova AS, Song RJ, Staunton D, Stoddard C, Stott K, Strauss HM, Streicher WW, Sumida JP, Swygert SG, Szczepanowski RH, Tessmer I, Toth RT, Tripathy A, Uchiyama S, Uebel SFW, Unzai S, Gruber AV, von Hippel PH, Wandrey C, Wang SH, Weitzel SE, Wielgus-Kutrowska B, Wolberger C, Wolff M, Wright E, Wu YS, Wubben JM, Schuck P

Publication type: Article

Publication status: Published

Journal: PLoS One

Year: 2015

Volume: 10

Issue: 5

Print publication date: 01/01/2015

Online publication date: 21/05/2015

Acceptance date: 02/04/2015

Date deposited: 20/03/2019

ISSN (electronic): 1932-6203

Publisher: Public Library of Science

URL: http://dx.doi.org/10.1371/journal.pone.0126420

DOI: 10.1371/journal.pone.0126420


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Funding

Funder referenceFunder name
A.N.U. Senior Research Fellowship
Instruct, European Strategy Forum on Research Infrastructures (ESFRI)
Intramural Research Programs of the National Institute of Biomedical Imaging and Bioengineering
National Heart, Lung, and Blood Institute
National Institute of Diabetes and Digestive and Kidney Diseases National Institutes of Health
NSF
EMBO long-term fellowship
Marie Curie IEF fellowship
Max Planck Society
2011-0010437National Research Foundation of Korea
ANR-10-INSB-05-01French Infrastructure for Integrated Structural Biology (FRISBI)
ANR-10-INSB-05-02FRISBI
ANR-10-LABX-49-01GRAL within the Grenoble Partnership for Structural Biology (PSB)
CA35635National Cancer Institute
CTQ2012-33717MINECO-Spain/FEDER
F31GM105363NIH
FZ 82Rudolf Virchow Center for Experimental Biomedicine - German Research Council (Deutsche Forschungsgemeinschaft, DFG)
GM 109102National Institute of General Medical Sciences
GM 094363National Institute of General Medical Sciences
GM 095822National Institute of General Medical Sciences
GM067777NIH
GM-15792NIH
GM-29158NIH
GM82545NIH
P30GM103519National Institute of General Medical Sciences
POIG.02.02.00-00025/09European Regional Development Fund within the Innovation Economy Operational Program
POIG.02.02.00-14-024/08-00Centre for Preclinical Research and Technology (CePT) - European Union

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