Non-Contact Universal Sample Presentation for Room Temperature Macromolecular Crystallography Using Acoustic Levitation

Abstract

Macromolecular Crystallography is a powerful and valuable technique to assess protein structures. Samples are commonly cryogenically cooled to minimise radiation damage effects from the X-ray beam, but low temperatures hinder normal protein functions and this procedure can introduce structural artefacts. Previous experiments utilising acoustic levitation for beamline science have focused on Langevin horns which deliver significant power to the confined droplet and are complex to set up accurately. In this work, the low power, portable TinyLev acoustic levitation system is used in combination with an approach to dispense and contain droplets, free of physical sample support to aid protein crystallography experiments. This method facilitates efficient X-ray data acquisition in ambient conditions compatible with dynamic studies. Levitated samples remain free of interference from fixed sample mounts, receive negligible heating, do not suffer significant evaporation and since the system occupies a small volume, can be readily installed at other light sources.

Towards an Optimal Sample Delivery Method for Serial Crystallography at XFEL

“…Being non-contact ADE has distinct advantages in liquid handling and has found great application throughout the crystallography pipeline starting from crystallization [150], microseeding [151], and ligand soaking [152][153][154] to crystal mounting [155,156]. Since droplets are ejected at a frequency compatible with the repetition rate of first generation XFEL facilities, ADE has also been utilized as a sample delivery method at both XFEL and synchrotron [157][158][159][160]. By altering the transducer's frequency, the volume of ejected droplets can be tuned to accommodate different sizes of crystals from 5 to 800 µm [160,161].…”

“…Since droplets are ejected at a frequency compatible with the repetition rate of first generation XFEL facilities, ADE has also been utilized as a sample delivery method at both XFEL and synchrotron [157][158][159][160]. By altering the transducer's frequency, the volume of ejected droplets can be tuned to accommodate different sizes of crystals from 5 to 800 µm [160,161]. Ejected droplets carry significant volumes of mother liquor [157], which contributes to more background compared to jet or fixed-target chip [157].…”

“…This requires very high accuracy and precision in the ejection trajectory and timing [157]. Alternatively, ejected droplets could be trapped in pressure node and levitated into mid-air using a reflector [159,160]. This maintains the position of the droplet while being probed by X-rays.…”

Beyond the Langevin horn: Transducer arrays for the acoustic levitation of liquid drops

“…In sample presentation scenarios however, the shape of the droplet can be an important factor which directly impacts the results. Consequently, there is an increasing shift from awareness towards control of droplet shape by balancing the confinement potential of the acoustic field against the forces applied to the droplets to maximise sphericity, for example in Non-Contact Universal Sample Presentation for Room Temperature Macromolecular Crystallography Using Acoustic Levitation [50] the droplet sphericity as a function of voltage applied to the TinyLev system was determined and optimised to balance these key parameters.…”

“…In recent publications such as [50] this new era of low-cost phased array ultrasonic levitation devices are beginning to find use in sample presentation to non-contact measurement techniques in applications previously making use of Langevin horns [51]. This facilitates containerless, background free spectroscopy which ushers in a new wave of experimental techniques and brings with it significant advantages in terms of measurement resolution without imparting significant energy into the sample.…”