Sample delivery for serial crystallography at free-electron lasers and synchrotrons

Abstract

The high peak brilliance and femtosecond pulse duration of X-ray free-electron lasers (XFELs) provide new scientific opportunities for experiments in physics, chemistry and biology. In structural biology, one of the major applications is serial femtosecond crystallography. The intense XFEL pulse results in the destruction of any exposed microcrystal, making serial data collection mandatory. This requires a high-throughput serial approach to sample delivery. To this end, a number of such sample-delivery techniques have been developed, some of which have been ported to synchrotron sources, where they allow convenient low-dose data collection at room temperature. Here, the current sample-delivery techniques used at XFEL and synchrotron sources are reviewed, with an emphasis on liquid injection and high-viscosity extrusion, including their application for time-resolved experiments. The challenges associated with sample delivery at megahertz repetition-rate XFELs are also outlined.

Shortening injection matrix for serial crystallography

“…In SX experiments, it is important to deliver the crystal sample serially and stably to the X-ray interaction point. 5 Delivering the sample reliably not only reduces sample consumption but also enables data collection for accurate responses in time-resolved studies. 6 Among the various sample delivery methods, including liquid jet injector, 7 high viscosity injector, 8,9 fixed-target scanning, 10–12 and microfluidics, 13,14 sample delivery using a viscous medium with an injector or syringe is widely applied to serial femtosecond crystallography (SFX) or serial millisecond crystallography (SMX), which can successfully determine structure using an XFEL or synchrotron, respectivley.…”

Shortening injection matrix for serial crystallography

“…Therefore, SX experiments allow for in-depth analyses of the more biologically reliable crystal structure of macromolecules beyond what is possible with conventional X-ray crystallography. In SX experiments, it is important to deliver the crystal sample serially and stably to the X-ray interaction point 10 . Delivering the sample reliably not only reduces sample consumption but also enables data collection for accurate responses in time-resolved studies 11 .…”

Viscous medium-based crystal support in sample holder for fixed-target serial femtosecond crystallography

“…SFX using an intense X-ray free electron laser (XFEL) with an ultrashort pulse can be used to analyze smaller crystals than those used in traditional X-ray crystallographic methods; however, this method requires a large number of crystal samples, as each crystal is exposed only once to the Xray beam (Boutet et al, 2012). Thus, this technique requires a delivery technology that serially and reliably delivers numerous crystal samples to the X-ray interaction position (Grunbein & Nass Kovacs, 2019). Various sample delivery methods such as injector-based methods (DePonte et al, 2008, Weierstall et al, 2014, syringe method (Sugahara et al, 2015, Berntsen et al, 2019, capillary method (Stellato et al, 2014, Nam, 2020a, fixed-target scanning (Hunter et al, 2014, Cohen et al, 2014, Mueller et al, 2015, Lee et al, 2019, electrospinning (Sierra et al, 2012), and microfluidic devices (Monteiro et al, 2019) have been used for crystal delivery in serial crystallography (SX) using XFEL or synchrotron X-rays.…”