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A Round-Robin Approach Provides a Detailed Assessment of Biomolecular Small-Angle Scattering Data Reproducibility and Yields Consensus Curves for Benchmarking.



Jill Trewhella, Patrice Vachette, Jan Bierma, Clement Blanchet, Emre Brookes, Srinivas Chakravarthy, Leonie Chatzimagas, Thomas Cleveland, Nathan Cowieson, Ben Crossett, Anthony P. Duff, Daniel Franke, Frank Gabel, Richard E. Gillilan, Melissa Graewert, Alexander Grishaev, Jules M. Guss, Michal Hammel, Jesse Hopkins, Qingqui Huang, Jochen S. Hub, Gregory L. Hura, Thomas C. Irving, Cy M. Jeffries, Cheol Jeong, Nigel Kirby, Susan N. Krueger, Anne Martel, Tsutomu Matsui, Na Li, Javier Perez, Lionel Porcar, Thierry Prange, Ivan Rajkovic, Mattia Rocco, Daniel J. Rosenberg, Timothy M. Ryan, Soenke Seifert, Hiroshi Sekiguchi, Dmitri Svergun, Susana C. Marujo Teixeira, Aurelien Thureau, Thomas M. Weiss, Andrew Whitten, Kathleen Wood, Xiaobing Zuo


Small-Angle Scattering (SAS) data from 5 candidate proteins (RNaseA, lysozyme, xylanase, urate oxidase and xylose isomerase) were measured on 12 Small-Angle X-ray Scattering (SAXS) and 4 Small-Angle Neutron Scattering (SANS) instruments. In total, more than 150 SAXS data sets and more than 70 SANS data sets were submitted for evaluation. Measurements in batch mode and with in-line Size Exclusion Chromatography (SEC) were made using SAXS and SANS, in H2O and D2O buffers. In-line SEC measurements proved generally superior to batch measurements for eliminating sample heterogeneity. Careful merging of data measured using SEC-SAS and batch SAS modes, or low- and high-concentration data from batch SAS measurements, was successful in eliminating small amounts of aggregate or interparticle interference from the scattering profile. In general, the experimental profiles were reproducible with the caveat that constant adjustment was required to account for small errors in solvent subtraction and dark current corrections. Consensus SAXS data sets over the q-measurement range 0 – 1 Å-1 for all 5 proteins showed good agreement with theoretical prediction. Additionally, 2 sets of data measured at different facilities (one using in-line SEC and the other batch mode) to q > 2.2 Å-1 showed good agreement indicating that this region has interpretable features. The inherently lower statistical precision for SANS limited the reliably measured q-range to 0.5 Å^-1^, but within the limits of experimental uncertainties there was also good agreement with prediction for the consensus SANS data for all 5 proteins measured in H2O and in D2O.
Acta Crystallographica Section D-Biological Crystallography


Scattering, SAXS, Small-Angle Neutron Scattering, SANS, scattering standards


Trewhella, J. , Vachette, P. , Bierma, J. , Blanchet, C. , Brookes, E. , Chakravarthy, S. , Chatzimagas, L. , Cleveland, T. , Cowieson, N. , Crossett, B. , Duff, A. , Franke, D. , Gabel, F. , Gillilan, R. , Graewert, M. , Grishaev, A. , Guss, J. , Hammel, M. , Hopkins, J. , Huang, Q. , Hub, J. , Hura, G. , Irving, T. , Jeffries, C. , Jeong, C. , Kirby, N. , Krueger, S. , Martel, A. , Matsui, T. , Li, N. , Perez, J. , Porcar, L. , Prange, T. , Rajkovic, I. , Rocco, M. , Rosenberg, D. , Ryan, T. , Seifert, S. , Sekiguchi, H. , Svergun, D. , Marujo Teixeira, S. , Thureau, A. , Weiss, T. , Whitten, A. , Wood, K. and Zuo, X. (2022), A Round-Robin Approach Provides a Detailed Assessment of Biomolecular Small-Angle Scattering Data Reproducibility and Yields Consensus Curves for Benchmarking., Acta Crystallographica Section D-Biological Crystallography, [online], (Accessed June 20, 2024)


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Created November 1, 2022, Updated January 23, 2024