Microfluidic approaches for the analysis of protein–protein interactions in solution

Abstract

Exploration and characterisation of the human proteome is a key objective enabling a heightened understanding of biological function, malfunction and pharmaceutical design. Since proteins typically exhibit their behaviour by binding to other proteins, the challenge of probing protein-protein interactions has been the focus of new and improved experimental approaches. Here, we review recently developed microfluidic techniques for the study and quantification of protein–protein interactions. We focus on methodologies that utilise the inherent strength of microfluidics for the control of mass transport on the micron scale, to facilitate surface and membrane-free interrogation and quantification of interacting proteins. Thus, the microfluidic tools described here provide the capability to yield insights on protein–protein interactions under physiological conditions. We first discuss the defining principles of microfluidics, and methods for the analysis of protein–protein interactions that utilise the diffusion-controlled mixing characteristic of fluids at the microscale. We then describe techniques that employ electrophoretic forces to manipulate and fractionate interacting protein systems for their biophysical characterisation, before discussing strategies that use microdroplet compartmentalisation for the analysis of protein interactions. We conclude by highlighting future directions for the field, such as the integration of microfluidic experiments into high-throughput workflows for the investigation of protein interaction networks.

Direct digital sensing of protein biomarkers in solution

“…Future iterations of the platform could also incorporate, for example, multicolor single-molecule spectroscopy and FRET techniques, [39][40][41] as well as other microfluidic separation modalities 42,43 combined with downstream analyses, 44 to enhance assay parallelization, sensitivity, and robustness to experimental noise.…”

In situ Electron Microscopy of Complex Biological and Nanoscale Systems: Challenges and Opportunities

“…The microfluidic design can be customized to allow controlled mixing of multiple input materials and to accommodate biological structures of different length-scales. For miscible liquids, the microchannel allows a continuous flow of analytes via diffusional mixing (Arter et al, 2020), i.e., achieved via passive mixing based on the physical design of the microchannel alone ( Figure 5B). Active mixing is possible by applying suitable external stimuli such as electric or magnetic fields, pressure, and acoustics (Solsona et al, 2019).…”

“…Strategies for creating such functional microchannels, by chemically or physically modifying the inner walls of the microchannels, have been summarized in a recent review (Wang et al, 2019). As the field progresses, additional functionalities of the microfluidic platform such as electrophoretic manipulation (Huh et al, 2008;Yasukawa et al, 2008;Unocic et al, 2014;Arter et al, 2020) can be incorporated for more advanced characterization.…”

“…This will facilitate the understanding of the operative mechanism of hydrotropes as more biological hydrotropes become discovered. It can have further implications in understanding protein-protein interactions, or interactomes (Tian et al, 2011;Arter et al, 2020), in vivo.…”

Potential of Microfluidics and Lab-on-Chip Platforms to Improve Understanding of “prion-like” Protein Assembly and Behavior

“…In a laminar flow regime mixing only occurs by diffusion. Thus, the mixing of the analytes depends only on their intrinsic diffusion coefficient (see Arter et al, 2020 for a recent review). After this, the diffusion parameters of the different elements (i.e., oligomers) can be controlled and analyzed during amyloid aggregation.…”