Magnetic Nanoparticle Tracking for One-Step Protein Separation and Binding Kinetics Analysis

Label-free techniques for quantification of protein-protein interaction often require protein samples separated from complex media using affinity purification tools such as magnetic nanoparticles. However, the separated proteins are attached to the nanoparticles and require additional preparation steps, including elution and immobilization to a sensor surface before measurement. To streamline this tedious process, we present a method that can directly quantify the protein binding kinetics on nanoparticles without elution and immobilization, by optically tracking the nanoparticle size change upon ligand binding. We measured antibody binding to nanoparticles with captured protein, which was pulled down from a different medium prior to the measurement. The source of noise for the method was also analyzed. We anticipate this method can simplify the workflow from protein separation to detection while providing sufficient binding kinetics and affinity information for protein studies.