Optical and electrochemical immunoassay for detection of superoxide dismutase 2 employing biocatalytic formation of quantum dots on the surface of microbeads

INTRODUCTION: Semiconductor nanocrystals known as quantum dots (QDs) are increasingly being used as photoluminescent materials in bio-imaging, photonics, and optoelectronic applications. Our group has pioneered the enzymatic... [ view full abstract ]

INTRODUCTION:
Semiconductor nanocrystals known as quantum dots (QDs) are increasingly being used as photoluminescent materials in bio-imaging, photonics, and optoelectronic applications. Our group has pioneered the enzymatic in situ growth1-4 and etching 5 of fluorescent semiconductor CdS QDs, and on this base developed fluorogenic enzymatic assays. Those assays were much more cost efficient and demonstrated lower detection limits and better sensitivities than the standard assays based on organic substrates. In this study we developed new optical and electrochemical immunoassay for detection of tumor biomarkers such as superoxide dismutase (SOD2) employing enzymatic in situ generation and immobilization of CdS QDs onto microspheres.

METHODS:
Cadmium sulfide semiconductor nanoparticles were detected by fluorescence spectroscopy and square-wave voltammetry. Characterized by a wide field fluorescence microscope and a transmission electron microscopy equipped with an Energy Dispersive X-Ray Spectrometer.

RESULTS AND DISCUSSION:
In our immunoassay (Sheme 1) the target analyte (A) SOD2 mediates immobilization of alkaline phosphatase antibody conjugate on the surface of polyvinyl chloride microbeads. The enzymatic hydrolysis of para-nitrophenylphosphate by alkaline phosphatase triggered rapid formation on the surface of microbeads of phosphate-stabilized cadmium sulfide semiconductor nanoparticles which were detected by fluorescence spectroscopy and square-wave voltammetry. We have demonstrated that electrochemical and fluorogenic detection employing enzymatically generated CdS NPs yield new immunoassays with better detection limits in comparison with those of the previously published methods at least by three orders of magnitude. Our methodology allows for the detection of SOD2 in lysates from HepG2 (Human hepatocellular carcinoma) cells.

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