Abstract
Serial femtosecond crystallography (SFX) uses X-ray pulses from free-electron laser (FEL) sources that can outrun radiation damage and thereby overcome long-standing limits in the structure determination of macromolecular crystals. Intense X-ray FEL pulses of sufficiently short duration allow the collection of damage-free data at room temperature and give the opportunity to study irreversible time-resolved events. SFX may open the way to determine the structure of biological molecules that fail to crystallize readily into large well-diffracting crystals. Taking advantage of FELs with high pulse repetition rates could lead to short measurement times of just minutes. Automated delivery of sample suspensions for SFX experiments could potentially give rise to a much higher rate of obtaining complete measurements than at today's third generation synchrotron radiation facilities, as no crystal alignment or complex robotic motions are required. This capability will also open up extensive time-resolved structural studies. New challenges arise from the resulting high rate of data collection, and in providing reliable sample delivery. Various developments for fully automated high-throughput SFX experiments are being considered for evaluation, including new implementations for a reliable yet flexible sample environment setup. Here, we review the different methods developed so far that best achieve sample delivery for X-ray FEL experiments and present some considerations towards the goal of high-throughput structure determination with X-ray FELs.
Highlights
Intense X-ray free-electron laser (FEL) pulses of sufficiently short duration allow the collection of damage-free data at room temperature and give the opportunity to study irreversible time-resolved events
Various sample delivery systems have been implemented for experiments at the Linac Coherent Light Source (LCLS) in California and the SPring-8 Angstrom Compact free-electron LAser (SACLA) in Japan
We review here different sample delivery methods developed for experimenting at X-ray FELs and describe concepts for a sample environment that could facilitate automation in data collection, sample screening, and nozzle exchange, which may allow SFX experiments to be carried out at high efficiency
Summary
Various approaches have been developed and tried to deliver large numbers of small crystals of micrometer size or smaller to the beam, for the purpose of measuring a single diffraction pattern per shot Under these conditions, there is no need to cryogenically cool the sample when using femtosecond X-ray pulses; samples can be measured at room temperature. Since the data are collected one grain at a time, it is possible to index each pattern and perform the summation on reflections of like Miller indices In this way, a set of three-dimensional structure factors are obtained, where each is averaged over the same set of crystal shapes and sizes and beam fluctuations that would have produced a (nonoverlapping) Debye-Scherrer ring. We review here different sample delivery methods developed for experimenting at X-ray FELs and describe concepts for a sample environment that could facilitate automation in data collection, sample screening, and nozzle exchange, which may allow SFX experiments to be carried out at high efficiency
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