Segmented flow generator for serial crystallography at the European X-ray free electron laser

Area of Science:

• Structural Biology
• Biophysics
• X-ray Science

Background:

• Serial femtosecond crystallography (SFX) using X-ray free electron lasers (XFELs) is crucial for determining structures of challenging targets like membrane proteins.
• Current continuous liquid sample delivery methods for SFX are highly inefficient due to the pulsed nature of XFELs, wasting over 99% of valuable protein crystal samples.

Purpose of the Study:

• To develop and demonstrate a novel microfluidic device for droplet-based sample delivery in SFX experiments.
• To reduce sample waste and enable efficient structure determination using XFELs.

Main Methods:

• A microfluidic device was designed to deliver protein crystals encapsulated in droplets, segmented by an immiscible oil.
• This droplet injection system was tested and validated at the European XFEL (EuXFEL) for high-pressure liquid SFX experiments.

Main Results:

• The microfluidic device achieved approximately a 60% reduction in sample waste compared to continuous jetting.
• The structure of 3-deoxy-D-manno-octulosonate-8-phosphate synthase was successfully determined from microcrystals delivered via droplets.
• Novel structural features of the enzyme were revealed through this new approach.

Conclusions:

• Microfluidic droplet-based sample delivery is a viable and significantly more efficient alternative for SFX experiments at XFELs.
• This technology advances the capability for time-resolved studies and structure determination of scarce or precious samples.