Formation and Characterization of Stable Antifouling Colloidal Particles via Surface-Initiated-Photoinduced Electron Transfer-Reversible Addition–Fragmentation Chain-Transfer Polymerization for Standardized Biophysical Measurements

Extracellular vesicles (EVs) are potential biomarkers for various diseases, but EV concentration measurements are difficult to achieve, which often leads to incomparable and irreproducible data. To solve this problem, colloidal particles of known concentration could assist in tracking the sample dilution and determining the volume of the sample in which EVs are counted during the analysis. However, colloidal particles in body fluids suffer from instability due to the nonspecific adsorption of proteins onto their surface. Our aim is to functionalize colloidal particles with antifouling polymer brushes to prevent nonspecific adsorption of proteins from body fluids. To functionalize polystyrene colloidal particles with a diameter of 220 nm, surface-initiated photoinduced electron transfer-reversible addition-fragmentation chain-transfer polymerization was used with three different monomers: nonzwitterionic <i>N</i>-(2-hydroxypropyl)methacrylamide (HPMA), zwitterionic methacrylate phosphocholine, and carboxybetaine methacrylate. Polymer brush growth was confirmed with dynamic light scattering, X-ray photoelectron spectroscopy, and transmission electron microscopy. The functionalized colloidal particles were exposed to a single-protein solution and human blood plasma, wherein HPMA-functionalized colloidal particles demonstrated excellent antifouling. Moreover, HPMA-functionalized colloidal particles were added to pure plasma and remained stable for at least 5 h. The successful functionalization of colloidal particles with antifouling polymer brushes shows great potential for further research in both flow cytometry and biomedical applications, particularly in selective biomarker detection.