A high-transparency, micro-patternable chip for X-ray diffraction analysis of microcrystals under native growth...

Area of Science:

• Structural Biology
• Biophysics
• Crystallography

Background:

• Microcrystals hinder macromolecular structure determination via X-ray diffraction.
• Microfocus synchrotron beamlines and X-ray free-electron lasers (XFELs) can collect data from microcrystals.
• Efficient methods are needed to deliver small volumes of microcrystals under native conditions for X-ray analysis.

Purpose of the Study:

• To design and present a novel micro-diffraction device for efficient microcrystal analysis.
• To enable rapid sample delivery and data collection from microcrystals at X-ray beam sources.
• To overcome challenges in analyzing microcrystals grown in common laboratory formats.

Main Methods:

• Fabrication of a three-layer micro-diffraction device using silicon nitride, photoresist, and polyimide film.
• Design incorporates X-ray-transparent materials to minimize background scattering.
• Customizable surface patterns (hydrophobic/hydrophilic) localize microcrystals, allowing loading via standard pipette.

Main Results:

• The device exhibits low X-ray scattering and absorption.
• High-resolution (<1.6 Å) diffraction data were collected from hen egg-white lysozyme microcrystals (10-15 µm).
• A complete dataset was sufficient for high-quality structure determination by molecular replacement.

Conclusions:

• The developed micro-diffraction chip facilitates user-friendly analysis of microcrystals.
• Enables rapid data collection at room temperature using microfocus synchrotron beamlines and XFELs.
• The device supports analysis of microcrystals grown in various laboratory formats, advancing structural biology.