Acoustic methods for high-throughput protein crystal mounting at next-generation macromolecular crystallographic beamlines

Abstract

To take full advantage of advanced data collection techniques and high beam flux at next-generation macromolecular crystallography beamlines, rapid and reliable methods will be needed to mount and align many samples per second. One approach is to use an acoustic ejector to eject crystal-containing droplets onto a solid X-ray transparent surface, which can then be positioned and rotated for data collection. Proof-of-concept experiments were conducted at the National Synchrotron Light Source on thermolysin crystals acoustically ejected onto a polyimide 'conveyor belt'. Small wedges of data were collected on each crystal, and a complete dataset was assembled from a well diffracting subset of these crystals. Future developments and implementation will focus on achieving ejection and translation of single droplets at a rate of over one hundred per second.

Precise Manipulation and Patterning of Protein Crystals for Macromolecular Crystallography Using Surface Acoustic Waves

“…The most successfully implemented method for manipulating crystals uses acoustic droplet ejection. [15] This method ejects small droplets containing crystals onto a surface. It requires an expensive, complex setup.…”

Solvent minimization induces preferential orientation and crystal clustering in serial micro-crystallography on micro-meshes,in situplates and on a movable crystal conveyor belt

“…When crystals were found, a 6 wedge of data was obtained (either a single 6 rotation or three 2 rotations). Serial data were obtained from crystals deposited on micromeshes (a), on in situ micro-plates (b) and on a moving conveyor belt system (c) (Roessler et al, 2013). In each case, diffraction from $100 individual crystals was collated into a high-quality data set.…”

“…1c). Data were obtained from crystals that were serially transferred to a goniometer-mounted conveyor belt system (Roessler et al, 2013).…”

“…Faced with the inevitable trade-offs between different capabilities, we were motivated to begin finding answers to some of these questions. Our group has successfully demonstrated unique new approaches to serial micro-crystallography, such as acoustic droplet ejection (Ellson et al, 2003), which uses sound waves to move crystals directly from their growth trays onto data-collection media (Soares et al, 2011;Roessler et al, 2013) or to grow protein crystals on plates (Villaseñ or et al, 2012) or directly on micro-meshes (Yin et al, 2014). Initially, our serial crystallography efforts were frustrated by difficulties such as preferential orientation and crystal clustering.…”

“…Serial micro-crystallography efforts currently explore diverse strategies for combining data from a large number of crystals that transit the X-ray beam either tumbling freely (Boutet & Robinson, 2008;Weierstall et al, 2008) or deposited on a surface (Ji et al, 2010;Soares et al, 2011;Roessler et al, 2013). New X-ray sources enable the use of micro-crystals because they can focus down to a few micrometers.…”