Target Confinement in Small Reaction Volumes Using Microfluidic Technologies: A Smart Approach for Single-Entity Detection and Analysis.

Over the last decades, the study of cells, nucleic acid molecules, and proteins has evolved from ensemble measurements to so-called single-entity studies. The latter offers huge benefits, not only as biological research tools to examine heterogeneities among individual entities within a population, but also as biosensing tools for medical diagnostics, which can reach the ultimate sensitivity by detecting single targets. Whereas various techniques for single-entity detection have been reported, this review focuses on microfluidic systems that physically confine single targets in small reaction volumes. We categorize these techniques as droplet-, microchamber-, and nanostructure-based and provide an overview of their implementation for studying single cells, nucleic acids, and proteins. We furthermore reflect on the advantages and limitations of these techniques and highlight future opportunities in the field.