High throughput protein nanocrystal fractionation in a microfluidic sorter

Area of Science:

• Biophysics
• Structural Biology
• Biotechnology

Background:

• Serial femtosecond crystallography (SFX) using X-ray free electron lasers (XFELs) is revolutionizing protein structure determination.
• High-quality diffraction data in SFX requires a narrow distribution of small protein crystal sizes.
• Current methods for obtaining such samples are often inefficient or lack precision.

Purpose of the Study:

• To optimize a microfluidic device for dielectrophoretic sorting of protein crystals.
• To enhance throughput and control over size-based fractionation for SFX applications.
• To achieve submicrometer size fractions of protein crystals suitable for advanced crystallography.

Main Methods:

• Computational modeling of applied potentials and device geometry for optimal sorting efficiency.
• Systematic optimization of critical geometrical dimensions and input parameters for nanoparticle sorting.
• Experimental validation using nanobeads and protein crystals to assess fractionation performance.

Main Results:

• The optimized microfluidic device demonstrated high sorting efficiencies for separating nanoparticles from microparticles.
• Successful fractionation of protein crystals into desired submicrometer size ranges was achieved.
• Improved throughput and control over the sorting process were validated experimentally.

Conclusions:

• The optimized microfluidic dielectrophoretic sorter is a promising tool for preparing protein crystal samples for SFX.
• This technology addresses a critical bottleneck in enabling high-resolution structural studies of complex proteins.
• Further development holds potential for broader applications in bioparticle separation and analysis.