Diffusion-based size determination of solute particles: a method adapted for PSD proteins

The postsynaptic density (PSD) is a complex, multi-layered protein network largely situated on the internal surface of the postsynaptic membrane, with receptor proteins extending it across the membrane. It is the first processing unit for incoming synaptic transmissions, and changes in its internal structure exhibit a strong correlation with synaptic strength and plasticity. These structural changes are largely governed by multivalent interactions between its components, leading to complex formation and, under certain circumstances, protein phase separation. Here we present an experimental method for detecting specific PSD proteins as well as their complexes via their diffusion. The method requires a fluorescent labelling technique that does not disrupt the function of labelled proteins, a microfluidic device that can maintain laminar flow for labelled protein solutions, a microscope that can record the fluorescent signal emitted by these solutions, and an analytic software package that can process the collected experimental data and convert them into approximate particle sizes.