Abstract
Reliable sample delivery is essential to biological imaging using X-ray Free Electron Lasers (XFELs). Continuous injection using the Gas Dynamic Virtual Nozzle (GDVN) has proven valuable, particularly for time-resolved studies. However, many important aspects of GDVN functionality have yet to be thoroughly understood and/or refined due to fabrication limitations. We report the application of 2-photon polymerization as a form of high-resolution 3D printing to fabricate high-fidelity GDVNs with submicron resolution. This technique allows rapid prototyping of a wide range of different types of nozzles from standard CAD drawings and optimization of crucial dimensions for optimal performance. Three nozzles were tested with pure water to determine general nozzle performance and reproducibility, with nearly reproducible off-axis jetting being the result. X-ray tomography and index matching were successfully used to evaluate the interior nozzle structures and identify the cause of off-axis jetting. Subsequent refinements to fabrication resulted in straight jetting. A performance test of printed nozzles at an XFEL provided high quality femtosecond diffraction patterns.
Highlights
1.1 Sample delivery in serial femtosecond crystallographyWith peak intensities that are 108 – 1010 times greater than those of synchrotron sources [1] and the ability to out-run radiation damage, the X-ray free electron laser (XFEL) has enabled novel methodological advances in protein crystallography using serial femtosecond crystallography (SFX) [2,3]
Sample delivery in serial femtosecond crystallography experiments (SFX) requires transporting millions of sensitive protein crystals of submicron dimensions, often in high vacuum, across the pathway of the pulsed X-ray beam. This can be achieved using a liquid jet formed in a Gas Dynamic Virtual Nozzle (GDVN), which focuses a liquid stream of a suitable buffer solution containing the crystals into the Xray beam
A 100-μm inner diameter capillary of approximately 2 meters in length was used for the gas supply line with a minimum operational gas pressure for the 3D printed nozzles of about 50 psi as measured upstream of the supply capillary (300 to 500 psi is typical of handmade GDVNs)
Summary
With peak intensities that are 108 – 1010 times greater than those of synchrotron sources [1] and the ability to out-run radiation damage, the X-ray free electron laser (XFEL) has enabled novel methodological advances in protein crystallography using serial femtosecond crystallography (SFX) [2,3]. Sample delivery in SFX requires transporting millions of sensitive protein crystals of submicron dimensions, often in high vacuum, across the pathway of the pulsed X-ray beam. This can be achieved using a liquid jet formed in a Gas Dynamic Virtual Nozzle (GDVN), which focuses a liquid stream of a suitable buffer solution containing the crystals into the Xray beam. Submicron jets down to about 0.3 microns in diameter have been demonstrated [7], which are of importance to the development of single-particle imaging methods This system has proven ideal for pump-probe time-resolved X-ray diffraction [4], and several promising schemes are under development to reduce protein consumption by eliminating the protein that runs to waste between X-ray pulses [8,9,10]. The accelerating liquid jet leaves the end of the outer glass capillary as a freely suspended continuous stream that subsequently breaks up into droplets due to the Rayleigh – Plateau instability [11] [Fig. 1]
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