Micropatterned assembly of silica nanoparticles for a protein microarray with enhanced detection sensitivity

We used an assembly of silica nanoparticles (SNPs) as a three-dimensional template for protein immobilization to prepare a protein microarray with enhanced protein loading capacity and detection sensitivity. SNPs were first modified with 3-aminopropyltriethoxysilane (APTES) for covalent immobilization of protein and micropatterned on poly(ethylene glycol)(PEG)-coated glass slides using elastomeric membranes with an array of holes. Proteins were selectively immobilized only on the SNP region, while the PEG regions served as an effective barrier to protein adsorption. Because of multi-layered SNPs that had curved surface, protein loading in the SNP micropattern was about six times greater than on a planar surface, as observed by fluorescence microscopy, which consequently improved the protein activity and reaction rate. GOX-catalyzed glucose oxidation and the molecular recognition mediated, specific binding between biotin and streptavidin were both successfully assayed using SNP microarrays, with better fluorescence signal and sensitivity than corresponding planar microarrays.