Co-flow injection for serial crystallography at X-ray free-electron lasers

Area of Science:

• Structural Biology
• Biophysics
• Materials Science

Background:

• Serial femtosecond crystallography (SFX) uses X-ray free-electron lasers for room-temperature macromolecular structure determination.
• Current SFX sample delivery methods, like liquid injection, often require large crystal quantities and can cause issues with vacuum compatibility and high background noise.
• These drawbacks limit the efficiency and scope of SFX experiments, particularly for time-resolved studies.

Purpose of the Study:

• To develop and characterize a novel sample delivery method for SFX that overcomes the limitations of existing techniques.
• To reduce protein crystal consumption and improve compatibility with beamline configurations.
• To enable advanced applications such as degradation-free light-induced time-resolved SFX.

Main Methods:

• A hybrid microfluidic 3D-printed co-flow device was designed and fabricated.
• An immiscible inert oil phase was used to supplement the sample flow in a co-flow configuration.
• Experimental characterization of co-flow generation with two resin materials was performed.
• A numerical model was developed to predict flow-rate conditions.

Main Results:

• Stable co-flow conditions were achieved with parallel sample and oil phases.
• The co-flow device demonstrated a significant reduction in crystal clogging.
• Potential for sample conservation up to 95% was shown.
• The method is compatible with vacuum configurations and reduces background noise.

Conclusions:

• The developed co-flow device offers a more efficient and sample-sparing method for SFX.
• This technique enhances the feasibility of time-resolved SFX experiments, including light-induced studies.
• The hybrid microfluidic approach represents a significant advancement in crystallographic sample delivery.