Microfluidics for Synchrotron to Unlock Structural Changes in Biomolecules

Microfluidics has emerged as a versatile platform for studying biomolecular processes and forming nanoparticles, due to its ability to manipulate fluids with precision at the microscale. This thesis reports the development of two microfluidic platforms: a stopped-flow system for time-resolved small-angle X-ray scattering (TR-SAXS) experiments at synchrotron beamlines, and a fast micromixer for lipid liquid crystalline nanoparticle (LLCN) formation.
The stopped-flow device integrates layered microfluidic chips, custom syringe drivers, and automated heating. A control system was developed and fully integrated with the EPICS/GDA environment at the I22 beamline of Diamond Light Source to make the device accessible to beamline users. Computational fluid dynamics (CFD) simulations guided optimisation of the vortex T-mixer geometry and operating conditions before fabrication. The device reduces sample requirements to 15 µl per experiment and achieves rapid and efficient mixing with a mixing index >0.95 and a dead time of 11 ms, validated using the reduction of 2,6-dichlorophenolindophenol (DCIP) reaction. Its performance was evaluated at synchrotron facilities, first by assessing mixing efficiency with an X-ray absorptive solution and then in time-resolved analyses to (i) track structural changes in nanoparticles undergoing cubic-to-hexagonal phase transitions and (ii) monitor the disruption of AdhE spirosomes by the anti-virulence compound ME0054, demonstrating its ability to capture structural dynamics across multiple timescales.
The fast mixer was developed for scalable and reproducible LLCN production, optimised through CFD simulations and fabricated using CNC machining. Its performance was tested against a commercial herringbone mixer, showing reliable formation of nanoparticles with controlled size and low polydispersity. Optimal flow conditions yielded cubosomes of ~170 nm with PDI values below 0.15, while SAXS confirmed preservation of the internal structure across operating ranges. The device matched, and in some cases outperformed, the herringbone mixer, demonstrating robustness and suitability for reproducible LLCN production.
Together, these developments provide open-source, beamline-compatible microfluidic tools that reduce sample consumption, improve reproducibility, and extend the scope of TR-SAXS experiments and nanoparticle production, contributing practical and accessible platforms for advancing research in soft matter and biomolecular science.