Construction of nanoparticle-based 3D protein microarrays with high protein capacity and controllable density

Abstract

Three-dimensional (3D) substrates have received considerable attention for the fabrication of protein microarrays with high protein capacity owing to their enlarged surface area for the high density immobilization of proteins. In this work, a facile strategy for constructing the protein microarrays with a low background and high signal intensity was developed based on the polystyrene nanoparticles (PS-NPs) assembled substrate. The discontinuous hydrophilic surface with hydrophobic initiators and hydrophilic poly(ethylene glycol) methacrylate (PEGMA) brushes was fabricated in the first step via digital micromirror device (DMD)-mediated photoinduced atom transfer radical polymerization (Photo-ATRP) process, and the hydrophilic interaction between sodium dodecyl sulfate (SDS)-modified PS-NPs and PEGMA brushes enabled the selective assembly of PS-NPs on the PEGMA brush region, resulting in the successful formation of arrayed surface with PS-NPs patterns. Then, poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC) brushes were grafted surrounding the PS-NPs to eliminate background noise caused by non-specific protein absorption. Compared to the PGMA brushes patterned substrate, the PS-NPs substrate exhibited excellent protein adsorption capability, leading to the 3D protein microarrays with higher protein capacity were easily obtained. Especially, the assembly density could be controlled by adjusting the PS suspension concentration, resulting in the effective regulation on the density of surface-bound proteins. Finally, the spatial control of protein density on the same substrate was achieved through regulating the distribution density of assembled PS-NPs, allowing for a protein microarrays with controllable density. This simple and effective mothed for fabricating nanoparticle-based 3D protein microarrays has tremendous potential in the fields of biomedical detection and high-throughput analysis.