Quantitative Real-Time Imaging of Protein-Protein Interactions by LSPR Detection with Micropatterned Gold Nanoparticles

Autor(en): Bhagawati, Maniraj
You, Changjiang 
Piehler, Jacob 
Stichwörter: AFFINITY; BINDING; Chemistry; Chemistry, Analytical; FUNCTIONAL IMMOBILIZATION; HISTIDINE-TAGGED PROTEINS; LABEL-FREE DETECTION; LAYERS; MASS-TRANSPORT; POLY(ETHYLENE GLYCOL); RECEPTOR INTERACTIONS; SURFACE-PLASMON RESONANCE
Erscheinungsdatum: 2013
Herausgeber: AMER CHEMICAL SOC
Journal: ANALYTICAL CHEMISTRY
Volumen: 85
Ausgabe: 20
Startseite: 9564
Seitenende: 9571
Zusammenfassung: 
Localized surface plasmon resonance (LSPR) offers powerful means for sensitive label-free detection of protein-protein interactions in a highly multiplexed format. We have here established self-assembly and surface modification of plasmonic nanostructures on solid support suitable for quantitative protein-protein interaction analysis by spectroscopic and microscopic LSPR detection. These architectures were obtained by layer-by-layer assembly via electrostatic attraction. Gold nanoparticles (AuNP) were adsorbed on a biocompatible amine-terminated poly(ethylene glycol) (PEG) polymer brush and further functionalized by poly-L-lysine graft PEG (PLL-PEG) copolymers. Stable yet reversible protein immobilization was achieved via tris(nitrilotriacetic acid) groups incorporated into the PLL-PEG coating. Thus, site-specific immobilization of His-tagged proteins via complexed Ni(II) ions was achieved. Functional protein immobilization on the surface was confirmed by real-time detection of LSPR scattering by reflectance spectroscopy. Association and dissociation rate constants obtained for a reversible protein-protein interaction were in good agreement with the data obtained by other surface-sensitive detection techniques. For spatially resolved detection, AuNP were assembled into micropatterns by means of photolithographic uncaging of surface amines. LSPR imaging of reversible protein-protein interactions was possible in a conventional wide field microscope, yielding detection limits of similar to 30 protein molecules within a diffraction-limited surface area.
ISSN: 00032700
DOI: 10.1021/ac401673e

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