title: Three-dimensional immobilization of beta-galactosidase on a silicon surface.
abstract: Many alternative strategies to immobilize and stabilize enzymes have been investigated in recent years for applications in biosensors. The entrapment of enzymes within silica-based nanospheres formed through silicification reactions provides high loading capacities for enzyme immobilization, resulting in high volumetric activity and enhanced mechanical stability. Here we report a strategy for chemically associating silica nanospheres containing entrapped enzyme to a silicon support. beta-galactosidase from E. coli was used as a model enzyme due to its versatility as a biosensor for lactose. The immobilization strategy resulted in a three-dimensional network of silica attached directly at the silicon surface, providing a significant increase in surface area and a corresponding 3.5-fold increase in enzyme loading compared to enzyme attached directly at the surface. The maximum activity recovered for a silicon square sample of 0.5 x 0.5 cm was 0.045 IU using the direct attachment of the enzyme through glutaraldehyde and 0.16 IU when using silica nanospheres. The immobilized beta-galactosidase prepared by silica deposition was stable and retained more than 80% of its initial activity after 10 days at 24 degrees C. The ability to generate three-dimensional structures with enhanced loading capacity for biosensing molecules offers the potential to substantially amplify biosensor sensitivity.
authors.affiliation: Air Force Research Laboratory, Tyndall AFB, Florida 32403, USA.
authors.forename: Lorena
authors.initials: L
authors.lastname: Betancor
authors.forename: Heather R
authors.initials: HR
authors.lastname: Luckarift
authors.forename: Jae H
authors.initials: JH
authors.lastname: Seo
authors.forename: Oliver
authors.initials: O
authors.lastname: Brand
authors.forename: Jim C
authors.initials: JC
authors.lastname: Spain
mesh_terms: D015374:Biosensing Techniques; D004800:Enzymes, Immobilized / Q000138:chemical synthesis*; D054118:Nanospheres; D012825:Silicon / Q000737:chemistry; D012822:Silicon Dioxide / Q000737:chemistry*; D013329:Structure-Activity Relationship; D001616:beta-Galactosidase / Q000737:chemistry* / Q000378:metabolism
chemical_list: D004800:Enzymes, Immobilized; D012822:Silicon Dioxide; D001616:beta-Galactosidase; D012825:Silicon