A user-friendly plug-and-play cyclic olefin copolymer-based microfluidic chip for room-temperature, fixed-target...

Area of Science:

• Structural Biology
• Biophysics
• Materials Science

Background:

• Serial X-ray crystallography advances protein structure determination, especially with X-ray free-electron lasers (XFELs).
• Efficient delivery of tiny, fragile protein crystals to the X-ray beam is critical for high-resolution data collection.
• Existing methods require improvements in speed, cost, and ease of use for diverse crystal types.

Purpose of the Study:

• To present an improved, all-polymer microfluidic chip design for room-temperature fixed-target serial crystallography.
• To create a user-friendly, adaptable, and cost-effective sample delivery platform for both synchrotron and XFEL facilities.
• To overcome limitations of previous designs, including cleanroom fabrication requirements.

Main Methods:

• Fabrication of an all-polymer microfluidic chip without cleanroom processes.
• Customization of chip design for various crystal types and sample loading methods (in situ or slurry).
• Validation using lysozyme and thaumatin crystals at synchrotron and XFEL beamlines.

Main Results:

• The improved chip design eliminates cleanroom fabrication, offering greater ease of use.
• Maintained crystal hydration stability with a larger imaging area to volume ratio.
• Successful data collection at synchrotron and XFEL facilities, achieving high-resolution structures (1.42–1.70 Å).

Conclusions:

• The novel microfluidic chip provides a robust, adaptable, and ultra-low-cost sample delivery solution for serial crystallography.
• Its simplified fabrication and modifiability make it readily adoptable for structural biologists, particularly with limited or fragile samples.
• This advancement facilitates high-resolution protein structure determination using X-ray crystallography.