Microtech2010 2010

Nanoscale Characterization of a Protease Dispersed Quantum Dot-Peptide Conjugate Assembly

J.G. Dick, P. Gryko, M. Ryan, M.M. Stevens
Imperial College London, UK

Keywords: quantum dots, gold nanoparticles, SAXS, uPA, TEM

Abstract:

A colloidal nanoparticle (NP)-based protease enzyme detection assay has been developed to detect urokinase-type Plasminogen Activator (uPA) - a prognostic biomarker for breast cancer. Rationally designed uPA substrate peptides were site-selectively conjugated to 1.4 nm gold NPs and immobilized to the surface of photoluminescent CdSe/ZnS quantum dots (QDs) via a hexahistidine amino acid sequence. The gold NPs tethered near the quantum dot surface quenched their luminescent signal by non-radiative energy transfer, analogous to FRET. Upon the introduction of the uPA protease, the peptide chains linking the gold NPs to the QD surface were cleaved, and the photoluminescence was recovered. The enzyme was detected in the nM concentration range. This system differs from commonly used ELISA protein detection assays by utilizing the activity of the enzyme to determine its concentration, and having the advantage of being quicker to perform without the need for expensive antibodies. This system was characterized by high resolution TEM at Imperial College London and small angle X-ray scattering (SAXS) at The Australian Synchrotron. These techniques facilitated the quantification of inter-particle spacing, stoichiometry and 3D structure of colloidal NP assemblies. This collaborative effort between Imperial College and The Australian Synchrotron allowed for a fastidious examination of a nanoparticle assembly to an extent not previously realized.
 
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