Multiscale Diffusion Measurements in Biological Gels Using Photoactivatable Fluorescent Nanoparticles

Abstract

Diffusion through biological gels is crucial for effective nanoparticle drug delivery. In this work, we develop a method to measure diffusivity over a large range of length scales -- from tens of nanometers to tens of microns -- using photoactivatable nanoparticle probes. Nanoparticles composed of block copolymers of poly(lactic-co-glycolic acid) (PLGA) and polyethylene glycol (PEG) were synthesized and confirmed to possess dense PEG coatings that resist bioadhesion, and these particles were conjugated with caged rhodamine to make them photoactivatable. Using confocal microscopy, we activated a region of these particles with a brief, targeted exposure to UV light, uncaging the rhodamine and causing the particles to become fluorescent. We observed their collective diffusion over tens of minutes and tens of microns, thus obtaining a measure of their diffusivity. This technique is complementary to traditional multiple particle tracking (MPT); it extends the range over which particle motion can be directly observed. We confirmed the accuracy of this technique with reference to MPT measurements and the known diffusivity of particles in water. We applied the method to a model fibrin gel system, and found that both our method and MPT measurements show an immobile fraction of particles and mobile fraction that diffuses over long scales. Finally, we examined nanoparticle diffusion in sputum collected from cystic fibrosis patients, and we measured particle diffusion over distances relevant to drug delivery in the lungs. Coupled with traditional MPT, this technique enables multiscale characterization of particle mobility in complex biological fluids