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).

The molecular level modification of surfaces: from self-assembled monolayers to complex molecular assemblies

“…73 The ability to stabilise PSi structures was vital for subsequent applications, where the photonic crystals were modified with peptides and used to detect the release of proteolytic enzymes from living cells. 131 The poor performance of the pre-synthesized molecule in the case of PSi photonic crystals was attributed to the oligo(ethylene oxide) moieties being highly flexible, and thus disrupting the packing of the SAM.…”

Wet chemical routes to the assembly of organic monolayers on silicon surfaces via the formation of Si–C bonds: surface preparation, passivation and functionalization

“…224 However, this one-pot approach toward the assembly of succinimidiyl ester-terminated surfaces requires significant synthetic effort as the relevant o-functionalized 1-alkene molecule is not commercially available and has to be customsynthesized. 58,59,225 We reason this as the major cause behind the limited number of literature examples on this directive approach, while on the other hand the step-wise NHS/EDC activation scheme (Si-RCOOH to Si-RC(O)NHS) remain widespread in surface science. 50,51,226,227 It is also worth noting that an optimization study to evaluate the optimal range of NHS and EDC concentrations and molar ratios has only very recently been published for porous silicon substrates by Sam et.…”

Vapor-phase silanization of oxidized porous silicon for stabilizing composition and photoluminescence

“…12,13 The devices fabricated with PSi have shown promise, owing to the ease of fabricating one-dimensional photonic crystals. 14 A significant benefit of biosensing with PSi photonic crystals is the labelfree transduction by optical interference ͑change in refractive index n within the film͒. 15,16 Unfortunately, the instability of the underlying surface to aqueous environments has hindered the wide-scale employment of PSi for biological applications.…”