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Review
. 2014 Jul 17;369(1647):20130337.
doi: 10.1098/rstb.2013.0337.

Liquid sample delivery techniques for serial femtosecond crystallography

Uwe Weierstall  1
Affiliations
Review

Liquid sample delivery techniques for serial femtosecond crystallography

Uwe Weierstall. Philos Trans R Soc Lond B Biol Sci. .

Abstract

X-ray free-electron lasers overcome the problem of radiation damage in protein crystallography and allow structure determination from micro- and nanocrystals at room temperature. To ensure that consecutive X-ray pulses do not probe previously exposed crystals, the sample needs to be replaced with the X-ray repetition rate, which ranges from 120 Hz at warm linac-based free-electron lasers to 1 MHz at superconducting linacs. Liquid injectors are therefore an essential part of a serial femtosecond crystallography experiment at an X-ray free-electron laser. Here, we compare different techniques of injecting microcrystals in solution into the pulsed X-ray beam in vacuum. Sample waste due to mismatch of the liquid flow rate to the X-ray repetition rate can be addressed through various techniques.

Keywords: X-ray free-electron laser; crystallography; liquid jets; serial femtosecond.

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Figures

Figure 1.
Figure 1.
Rayleigh jet issued from a 8 μm diameter nozzle. Rayleigh–Plateau instability causes breakup of the jet into droplets. The droplet break-up is triggered by a piezo transducer mounted to the nozzle, which is oscillating at 3.3 MHz.
Figure 2.
Figure 2.
GDVN nozzles as used at the LCLS, the arrow indicates the flow direction. (a) Nozzle is mounted in a stainless steel nozzle holder, which allows adjustment of the position of the inner capillary relative to the gas-focusing aperture. These nozzles can be disassembled and repaired if clogging occurs. (b) Glued nozzle, the inner capillary is glued into the outer tube. No repair or adjustment is possible. The length of the nozzle is about 5 cm.
Figure 3.
Figure 3.
Sucrose solution containing photosystem I crystals is injected into vacuum at a low flow rate of 300 nl min−1. The liquid capillary of 50 μm inner diameter protrudes out of the gas aperture. Ambient pressure acts on the liquid, and helium gas at 170 psi supply pressure is used as the coflowing gas. The liquid jet diameter can be reduced from the initial inner diameter of the liquid capillary to about 20 μm by the coflowing gas.
Figure 4.
Figure 4.
LCP injection into vacuum: a frame from a movie taken with the in vacuum camera during measurements at the LCLS. The nozzle shadow is on the right, and the X-ray pulse impact on the LCP jet (arrow) can be seen as a black bubble. LCP is extruded at a flow rate of 300 pl min−1. The pressure on the LCP was 3000 psi. The LCP extrusion speed was so slow that the X-ray pulse repetition rate at the LCLS had to be reduced to 1 Hz. X-ray energy: 9.4 keV, X-ray beam attenuated to 7%. LCP stream diameter: 20 μm.
Figure 5.
Figure 5.
DOD nozzle producing 30 μm diameter droplets, image taken with synchronized 100 ns flash exposures from an LED, repetition rate of droplet ejection was 120 droplets s−1 into ambient air.
Figure 6.
Figure 6.
Switch on time measurement of a pulsed GDVN nozzle. Two images from a high-speed camera movie recorded at 30 000 frames s−1. The liquid is turned on with a fast valve and the time difference between (a) and (b) is 367 µs. The gas flow is uninterrupted, only the liquid is pulsed. The time is shown on the upper right in milliseconds. The apparent offset of the stream from the centre in (b) is due to refraction in the outer glass tube.
Figure 7.
Figure 7.
GDVN nozzle operating in dripping mode, periodically ejecting liquid filaments with trailing droplets. (a–f) A sequence of images from a movie, taken during the ejection of one liquid filament, imaged with stroboscopic laser diode illumination. The liquid filament is initially compact (a–c), and then breaks up into droplet at the trailing end (d,e).
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