Facile strategy for protein conjugation with chitosan-poly(ethylene glycol) hybrid microparticle platforms via strain-promoted alkyne-azide cycloaddition (SPAAC) reaction

We demonstrate a facile fabrication-conjugation scheme for protein-conjugated biosensing platforms. Specifically, we utilize a chitosan-poly(ethylene glycol) hybrid system to fabricate highly uniform and chemically reactive microparticle platforms via simple replica molding. Strain-promoted alkyne-azide cycloaddition (SPAAC) reaction between azide-modified proteins and microparticles activated with strain-promoted cyclooctynes allows tunable protein conjugation under mild reaction conditions. Upon conjugation of a model red fluorescent protein, fluorescence and confocal micrographs show selective protein conjugation near the particle surfaces as well as long-term stability of the conjugation scheme. Fluorescence and AFM results upon conjugation with varying protein concentrations indicate controllable protein conjugation. Examination of protein-particle conjugation kinetics shows multiple reaction regimes; rapid initial, intermediate, and steady final stage. Lastly, we demonstrate antibody conjugation with the particles and selective and rapid target protein capture with antibody-conjugated particles. Combined, these results illustrate a facile fabrication-conjugation scheme for robust protein-conjugated platforms that can be readily enlisted in various protein sensing applications.