Peptide-modified optical filters for detecting protease activity.
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).
Kilian
Kristopher A
KA
School of Chemistry, Centre for Vascular Research, University of New South Wales, Sydney NSW 2052, Australia.
Böcking
Till
T
Gaus
Katharina
K
Gal
Michael
M
Gooding
J Justin
JJ
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
D055767:Immobilized Proteins; D010455:Peptides; D010447:Peptide Hydrolases; D012825:Silicon