Optical bunching of particles in a liquid flow

Abstract

High-speed liquid micro-jets are used to rapidly and repeatedly deliver protein microcrystals to focused and pulsed X-ray beams in the method of serial femtosecond crystallography. However, the current continuous flow of crystals is mismatched to the arrival of X-ray pulses, wasting vast amounts of an often rare and precious sample. Here, we introduce a method to address this problem by periodically trapping and releasing crystals in the liquid flow, creating locally concentrated crystal bunches, using an optical trap integrated in the microfluidic supply line. We experimentally demonstrate a 30-fold increase of particle concentration into 10 Hz bunches of 6.4 μm diameter polystyrene particles. Furthermore, using particle trajectory simulations, a comprehensive description of the optical bunching process and parameter space is presented. Adding this compact optofluidics device to existing injection systems would thereby dramatically reduce sample consumption and extend the application of serial crystallography to a greater range of protein crystal systems that cannot be produced in high abundance. Our approach is suitable for other microfluidic systems that require synchronous measurements of flowing objects.

Potential of Time-Resolved Serial Femtosecond Crystallography Using High Repetition Rate XFEL Sources

“…Future methods also need to be compatible with sub-micrometer crystals, i.e., they must exhibit a low solvent background. Most of these rely on the synchronization of the sample delivery with the arrival of the X-ray pulse either by bunching the crystals (optical bunching [46]), injecting an alternative liquid during the dark time (segmented flow droplet injection [45]) or aiming to reduce the sample amount needed for a stable jet [38,44] (see Figure 1).…”

“…The optical bunching method (Figure 1e) relies on the use of a laser setup to create an optical trap in the capillary to periodically trap and release concentrated crystals [46]. In the first description of this method, Awel et al demonstrated a 30-fold local sample increase in the concentration into 10 Hz bunches using polystyrene particles.…”

“…Figure 1. Schematic representations of standard (a,b) and innovative (c-e) injection delivery methods that are compatible with megahertz operation at the EuXFEL: (a) a GDVN-based liquid jet; (b) a DFFN-based liquid jet[38]; (c) a co-flow injection[44]; (d) a segmented flow droplet injection[45]; (e) a GDVN-based liquid jet with optical bunching[46].…”

Acoustic Focusing of Protein Crystals for In-Line Monitoring and Up-Concentration during Serial Crystallography

“… 16 , 17 Many of these nozzles were traditionally made by hand, but recent developments in 3D-printed nozzles 11 , 18 have increased the reproducibility and feasibility to integrate microfluidics that enable, for example, chemical reaction initiation through mix-and-inject SFX 19 or crystal bunching using optical traps. 20 Despite significant development, sample delivery is still often the major bottleneck at SFX experiments. Here, we address some of these challenges, specifically those due to lack of in-line diagnostics, concentration control, and potential leaks and clogs at unions and nozzles.…”

“…Many approaches have been developed to reliably deliver protein crystals to the interaction region, including gas-dynamic virtual nozzles, , double-flow focusing nozzles, , lipidic cubic phase injectors, drop-on-demand injectors, , high-speed fixed-target systems, and electrospun jets. , Many of these nozzles were traditionally made by hand, but recent developments in 3D-printed nozzles , have increased the reproducibility and feasibility to integrate microfluidics that enable, for example, chemical reaction initiation through mix-and-inject SFX or crystal bunching using optical traps . Despite significant development, sample delivery is still often the major bottleneck at SFX experiments.…”