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Article

Adding Size Exclusion Chromatography (SEC) and Light Scattering (LS) Devices to Obtain High-Quality Small Angle X-Ray Scattering (SAXS) Data

The European Molecular Biology Laboratory (EMBL) Hamburg, Notkestr. 85, 22607 Hamburg, Germany
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Crystals 2020, 10(11), 975; https://doi.org/10.3390/cryst10110975
Received: 1 October 2020 / Revised: 23 October 2020 / Accepted: 26 October 2020 / Published: 27 October 2020
We describe the updated size-exclusion chromatography small angle X-ray scattering (SEC-SAXS) set-up used at the P12 bioSAXS beam line of the European Molecular Biology Laboratory (EMBL) at the PETRAIII synchrotron, DESY Hamburg (Germany). The addition of size exclusion chromatography (SEC) directly on-line to the SAXS capillary has become a well-established approach to reduce the effects of the sample heterogeneity on the SAXS measurements. The additional use of multi-angle laser light scattering (MALLS), UV absorption spectroscopy, refractive index (RI), and quasi-elastic light scattering (QELS) in parallel to the SAXS measurements enables independent molecular weight validation and hydrodynamic radius estimates. This allows one to address sample monodispersity as well as conformational heterogeneity. The benefits of the current SEC-SAXS set-up are demonstrated on a set of selected standard proteins. The processed SEC-SAXS data and models are provided in the Small Angle Scattering Biological Data Bank (SASBDB) and are labeled as “bench-marked” datasets that include the unsubtracted data frames spanning the respective SEC elution profiles and corresponding MALLS-UV-RI-QELS data. These entries provide method developers with datasets suitable for testing purposes, in addition to an educational resource for SAS data analysis and modeling. View Full-Text
Keywords: small angle X-ray scattering; size exclusion chromatography; laser light scattering; data quality; benchmarked datasets; SASBDB small angle X-ray scattering; size exclusion chromatography; laser light scattering; data quality; benchmarked datasets; SASBDB
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MDPI and ACS Style

Graewert, M.A.; Da Vela, S.; Gräwert, T.W.; Molodenskiy, D.S.; Blanchet, C.E.; Svergun, D.I.; Jeffries, C.M. Adding Size Exclusion Chromatography (SEC) and Light Scattering (LS) Devices to Obtain High-Quality Small Angle X-Ray Scattering (SAXS) Data. Crystals 2020, 10, 975. https://doi.org/10.3390/cryst10110975

AMA Style

Graewert MA, Da Vela S, Gräwert TW, Molodenskiy DS, Blanchet CE, Svergun DI, Jeffries CM. Adding Size Exclusion Chromatography (SEC) and Light Scattering (LS) Devices to Obtain High-Quality Small Angle X-Ray Scattering (SAXS) Data. Crystals. 2020; 10(11):975. https://doi.org/10.3390/cryst10110975

Chicago/Turabian Style

Graewert, Melissa A., Stefano Da Vela, Tobias W. Gräwert, Dmitry S. Molodenskiy, Clément E. Blanchet, Dmitri I. Svergun, and Cy M. Jeffries. 2020. "Adding Size Exclusion Chromatography (SEC) and Light Scattering (LS) Devices to Obtain High-Quality Small Angle X-Ray Scattering (SAXS) Data" Crystals 10, no. 11: 975. https://doi.org/10.3390/cryst10110975

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