Falke Sven, Dierks Karsten, Blanchet Clement, Graewert Melissa, Cipriani Florent, Meijers Rob, Svergun Dmitri, Betzel Christian
Laboratory for Structural Biology of Infection and Inflammation, University Hamburg, c/o DESY, Building 22a, Notkestrasse 85, Hamburg 22603, Germany.
Xtal Concepts GmbH, Marlowring 19, Hamburg 22525, Germany.
J Synchrotron Radiat. 2018 Mar 1;25(Pt 2):361-372. doi: 10.1107/S1600577517017568. Epub 2018 Feb 13.
Small-angle X-ray scattering (SAXS) analysis of biomolecules is increasingly common with a constantly high demand for comprehensive and efficient sample quality control prior to SAXS experiments. As monodisperse sample suspensions are desirable for SAXS experiments, latest dynamic light scattering (DLS) techniques are most suited to obtain non-invasive and rapid information about the particle size distribution of molecules in solution. A multi-receiver four-channel DLS system was designed and adapted at the BioSAXS endstation of the EMBL beamline P12 at PETRA III (DESY, Hamburg, Germany). The system allows the collection of DLS data within round-shaped sample capillaries used at beamline P12. Data obtained provide information about the hydrodynamic radius of biological particles in solution and dispersity of the solution. DLS data can be collected directly prior to and during an X-ray exposure. To match the short X-ray exposure times of around 1 s for 20 exposures at P12, the DLS data collection periods that have been used up to now of 20 s or commonly more were substantially reduced, using a novel multi-channel approach collecting DLS data sets in the SAXS sample capillary at four different neighbouring sample volume positions in parallel. The setup allows online scoring of sample solutions applied for SAXS experiments, supports SAXS data evaluation and for example indicates local inhomogeneities in a sample solution in a time-efficient manner. Biological macromolecules with different molecular weights were applied to test the system and obtain information about the performance. All measured hydrodynamic radii are in good agreement with DLS results obtained by employing a standard cuvette instrument. Moreover, applying the new multi-channel DLS setup, a reliable radius determination of sample solutions in flow, at flow rates normally used for size-exclusion chromatography-SAXS experiments, and at higher flow rates, was verified as well. This study also shows and confirms that the newly designed sample compartment with attached DLS instrumentation does not disturb SAXS measurements.
生物分子的小角X射线散射（SAXS）分析越来越普遍，在SAXS实验之前，对全面、高效的样品质量控制的需求一直很高。由于SAXS实验需要单分散的样品悬浮液，最新的动态光散射（DLS）技术最适合获取溶液中分子粒径分布的非侵入性快速信息。在德国汉堡DESY的PETRA III的EMBL光束线P12的BioSAXS终端站设计并改装了一个多接收器四通道DLS系统。该系统能够在光束线P12使用的圆形样品毛细管内收集DLS数据。获得的数据提供了溶液中生物颗粒的流体动力学半径和溶液分散度的信息。DLS数据可以在X射线曝光之前和曝光期间直接收集。为了匹配P12处20次曝光约1秒的短X射线曝光时间，采用一种新颖的多通道方法，在SAXS样品毛细管中的四个不同相邻样品体积位置并行收集DLS数据集，大幅减少了迄今为止使用的20秒或通常更长的DLS数据收集时间。该装置允许对用于SAXS实验的样品溶液进行在线评分，支持SAXS数据评估，并例如以高效的方式指示样品溶液中的局部不均匀性。应用不同分子量的生物大分子来测试该系统并获取有关性能的信息。所有测量的流体动力学半径与使用标准比色皿仪器获得的DLS结果高度一致。此外，应用新的多通道DLS装置，还验证了在尺寸排阻色谱 - SAXS实验通常使用的流速以及更高流速下对流动样品溶液进行可靠的半径测定。这项研究还表明并证实，新设计的带有连接DLS仪器的样品室不会干扰SAXS测量。
