title: Peptide-modified optical filters for detecting protease activity.
abstract: The organic derivatization of silicon-based nanoporous photonic crystals is presented as a method to immobilize peptides for the detection of protease enzymes in solution. A narrow-line-width rugate filter, a one-dimensional photonic crystal, is fabricated that exhibits a high-reflectivity optical resonance that is sensitive to small changes in the refractive index at the pore walls. To immobilize peptide in the pore of the photonic crystal, the hydrogen-terminated silicon surface was first modified with the alkene 10-succinimidyl undecenoate via hydrosilylation. The monolayer with the succinimide ester moiety at the distal end served the dual function of protecting the underlying silicon from oxidation as well as providing a surface suitable for subsequent derivatization with amines. The surface was further modified with 1-aminohexa(ethylene glycol) (EG(6)) to resist nonspecific adsorption of proteins common in complex biological samples. The distal hydroxyl of the EG(6) is activated using the solid-phase coupling reagent disuccinimidyl carbonate for selective immobilization of peptides as protease recognition elements. X-ray photoelectron spectroscopy analysis reveals high activation and coupling efficiency at each stage of the functionalization. Exposure of the peptide-modified crystals to the protease subtilisin in solution causes a change in the refractive index, resulting in a shift of the resonance to shorter wavelengths, indicating cleavage of organic material within the pores. The lowest detected concentration of enzyme was 37 nM (7.4 pmol in 200 microL).
authors.affiliation: School of Chemistry, Centre for Vascular Research, University of New South Wales, Sydney NSW 2052, Australia.
authors.forename: Kristopher A
authors.initials: KA
authors.lastname: Kilian
authors.forename: Till
authors.initials: T
authors.lastname: Böcking
authors.forename: Katharina
authors.initials: K
authors.lastname: Gaus
authors.forename: Michael
authors.initials: M
authors.lastname: Gal
authors.forename: J Justin
authors.initials: JJ
authors.lastname: Gooding
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D055767:Immobilized Proteins / Q000737:chemistry*; D055095:Optics and Photonics / Q000295:instrumentation*; D010084:Oxidation-Reduction; D010447:Peptide Hydrolases / Q000032:analysis* / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry*; D016062:Porosity; D012825:Silicon / Q000737:chemistry; D013057:Spectrum Analysis
chemical_list: D055767:Immobilized Proteins; D010455:Peptides; D010447:Peptide Hydrolases; D012825:Silicon