In this paper in Analyst, Dr. Kiefer O. Ramberg, Prof. Martin Caffrey and co-authors demonstrate Flow Induced Dispersion Analysis (FIDA) as, in their words, a fast and simple method for characterising the association of a membrane protein in a detergent micelle with a soluble protein binder.

They set out the context as follows: membrane proteins are the most intensively studied targets in drug discovery; nanobodies raised against them have been formulated as therapeutics and used in structural biology as crystallisation chaperones and as fiducials for cryo-electron microscopy; and both applications call for stable complexes with dissociation constants in the nanomolar to picomolar range.

The authors list surface plasmon resonance, bio-layer interferometry and isothermal titration calorimetry as techniques that have been used for this purpose, and write of the comparison in their own words: "ITC measurements are generally slower and require significantly more sample, while providing a more comprehensive thermodynamic characterization including binding affinity, stoichiometry, and binding enthalpy. In comparison, immobilization-based platforms can require substantial assay development and surface-functionalization steps."

They describe the application of FIDA to membrane proteins as underexplored to date. In their experiments, sequential addition of the micellarised transporter protein to the labelled nanobody in detergent-containing buffer revealed single-digit nanomolar binding, which they report as consistent with binding constants obtained previously by surface plasmon resonance analysis. They describe the technique as efficient in sample consumption, stating that micrograms of protein in nanolitre volumes are required for full ligand titrations, and in speed of setup and data collection, with experiments on the order of minutes. Because FIDA is non-destructive and does not require immobilisation, they consider it feasible that samples may be reused, for example by preparing complexes on grids for structural characterisation by cryo-electron microscopy.

The authors propose that a protocol of this kind would expedite the development of novel nanobody-based biomedicines and facilitate nanobody-assisted membrane protein structural biology. It is an example of using FIDA to measure nanobody binding affinity and complex size for a membrane protein target in solution.