Light-induced Extracellular Vesicle Adsorption.

The role of extracellular vesicles (EVs) in human health and disease has garnered considerable attention over the past two decades. However, while several types of EVs are known to interact dynamically with the extracellular matrix and there is great potential value in producing high-fidelity EV micropatterns, there are currently no label-free, high-resolution, and tunable platform technologies with this capability. We introduce Light-induced Extracellular Vesicle Adsorption (LEVA) as a powerful solution to rapidly advance the study of matrix- and surface-bound EVs and other particles. The versatility of LEVA is demonstrated using commercial GFP-EV standards, EVs from glioblastoma bioreactors, and E. coli outer membrane vesicles (OMVs), with the resulting patterns used for single EV characterization, single cell migration on migrasome-mimetic trails, and OMV-mediated neutrophil swarming. LEVA will enable rapid advancements in the study of matrix- and surface-bound EVs and other particles, and should encourage researchers from many disciplines to create novel diagnostic, biomimetic, immunoengineering, and therapeutic screening assays.

Limiting Brownian Motion to Enhance Immunogold Phenotyping and Superimpose Optical and Non-Optical Single-EP Analyses.

Optical and non-optical techniques propelled the field of single extracellular particle (EP) research through phenotypic and morphological analyses, revealing the similarities, differences, and co-isolation of EP subpopulations. Overcoming the challenges of optical and non-optical techniques motivates the use of orthogonal techniques while analyzing extracellular particles (EPs), which require varying concentrations and preparations. Herein, we introduce the nano-positioning matrix (NPMx) technique capable of superimposing optical and non-optical modalities for a single-EP orthogonal analysis. The NPMx technique is realized by ultraviolet-mediated micropatterning to reduce the stochasticity of Brownian motion. While providing a systematic orthogonal measurement of a single EP via total internal reflection fluorescence microscopy and transmission electron microscopy, the NPMx technique is compatible with low-yield samples and can be utilized for non-biased electrostatic capture and enhanced positive immunogold sorting. The success of the NPMx technique thus provides a novel platform by marrying already trusted optical and non-optical techniques at a single-EP resolution.