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Sample Preparation and Data Collection for High-Speed Fixed-Target Serial Femtosecond Crystallography

Alke Meents, Philip Roedig, Helen M. Ginn, Tim Pakendorf, Geoff Sutton, Karl Harlos, Thomas S. Walter, Jan Meyer, Pontus Fischer, Ramona Duman, Ismo Vartiainen, Bernd Reime, Martin Warmer, Aaron S. Brewster, Iris D. Young, Tara Michels Clark, Nicholas K. Sauter, Marcin Sikorsky, Silke Nelson, Daniel S. Damiani, Roberto Alonso-Mori, Jingshan Ren, Elizabeth E. Fry, Christian David, David I. Stuart, Armin H. Wagner

Abstract

We have developed a protocol for high-speed fixed-target serial femtosecond crystallography (SFX) at X-ray free electron lasers (XFELs). Using the Roadrunner goniometer for fast scanning of microcrystals loaded on a micro-patterned silicon chip, our method allows for usage of the full 120 Hz repetition rate of the Linear Coherent Light Source (LCLS). By synchronizing the sample exchange to the arrival of the LCLS X-ray pulses our approach results in most efficient use of sample material and beamtime. The presented protocol describes the loading of microcrystals on micro-patterned silicon chips and subsequent data collection using the Roadrunner goniometer installed at an XFEL beamline. Data collection can be performed either at cryogenic or room temperature. We further describe the Roadrunner control and data acquisition software and data processing using cppxfel. Due to the small sample amounts required and the low background scattering levels achievable, the method is ideally suited for data collection of precious microcrystals from viruses or membrane proteins, where often only limited amounts are available. This protocol accompanies Roedig P et al, Nature Methods (10.1038/nmeth.4335, published online June 19, 2017).

Keywords
fixed-target serial femtosecond crystallography, protein crystallography, virus crystallography

Figures

Introduction

Equipment

Procedure

Timing

Troubleshooting

Anticipated Results

References

Acknowledgements

Supplementary Files

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Preprint: Please note that this article has not completed peer review.

Sample Preparation and Data Collection for High-Speed Fixed-Target Serial Femtosecond Crystallography

Alke Meents, Philip Roedig, Helen M. Ginn, Tim Pakendorf, Geoff Sutton, Karl Harlos, Thomas S. Walter, Jan Meyer, Pontus Fischer, Ramona Duman, Ismo Vartiainen, Bernd Reime, Martin Warmer, Aaron S. Brewster, Iris D. Young, Tara Michels Clark, Nicholas K. Sauter, Marcin Sikorsky, Silke Nelson, Daniel S. Damiani, Roberto Alonso-Mori, Jingshan Ren, Elizabeth E. Fry, Christian David, David I. Stuart, Armin H. Wagner

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Abstract

We have developed a protocol for high-speed fixed-target serial femtosecond crystallography (SFX) at X-ray free electron lasers (XFELs). Using the Roadrunner goniometer for fast scanning of microcrystals loaded on a micro-patterned silicon chip, our method allows for usage of the full 120 Hz repetition rate of the Linear Coherent Light Source (LCLS). By synchronizing the sample exchange to the arrival of the LCLS X-ray pulses our approach results in most efficient use of sample material and beamtime. The presented protocol describes the loading of microcrystals on micro-patterned silicon chips and subsequent data collection using the Roadrunner goniometer installed at an XFEL beamline. Data collection can be performed either at cryogenic or room temperature. We further describe the Roadrunner control and data acquisition software and data processing using cppxfel. Due to the small sample amounts required and the low background scattering levels achievable, the method is ideally suited for data collection of precious microcrystals from viruses or membrane proteins, where often only limited amounts are available. This protocol accompanies Roedig P et al, Nature Methods (10.1038/nmeth.4335, published online June 19, 2017).

Figures

Introduction

Equipment

Procedure

Timing

Troubleshooting

Anticipated Results

References

Acknowledgements

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