Three-Dimensional Optical and Computational Reconstruction of Colloidal Electrokinetic Flows via Multiplane Imaging and Multiscale Modeling.

Sorting, filtering, and controlling colloidal particles is crucial in many fields, ranging from chemistry to biology and physics. Dielectrophoresis (DEP) enables selective manipulation of colloids by non-uniform AC electric fields, yet the associated three-dimensional nanoparticle dynamics and electrokinetic flows remain difficult to access experimentally. Here, we combine real-time multiplane widefield microscopy (> 100 fps), 3D single-particle tracking, and multiscale modeling to reconstruct the motion of 200 nm fluorescent polystyrene nanoparticles around a quadrupolar electrode under positive and negative DEP. This optical-computational approach provides volumetric trajectories and flow maps, revealing particle trapping, depletion, and three-dimensional vortical motion. A model including DEP and AC electro-osmotic flow quantitatively reproduces the positive DEP dynamics, demonstrating direct agreement between experiment and theory. Under negative DEP, the model captures particle depletion near the electrodes but not all observed flow structures, pointing to additional electrokinetic or electrothermal contributions. This framework provides a route to validate and refine AC electrokinetic models for complex colloidal and microfluidic systems.