title: Microbead QD-ELISA: Microbead ELISA Using Biocatalytic Formation of Quantum Dots for Ultra High Sensitive Optical and Electrochemical Detection.
abstract: Electrochemical detection strategies employing semiconductor quantum dots (QDs) open up new opportunities for highly sensitive detection of biological targets. We designed a new assay based on microbead linked enzymatic generation of CdS QDs (Microbead QD-ELISA) and employed it in optical and electrochemical affinity assays for the cancer biomarker superoxide dismutase 2 (SOD2). Biotinylated antibodies against SOD2 were immobilized on the surface of polyvinyl chloride microbeads bearing streptavidin. In order to prevent any non-specific adsorption the microbeads were further blocked with bovine serum albumin. The analyte, SOD2 was captured on microbeads and labeled with alkaline phosphatase-conjugated antibody linked with mouse antibody against SOD2. Hydrolysis of para-nitrophenylphosphate by immobilized alkaline phosphatase triggered the rapid formation of phosphate-stabilized CdS QDs on the surface of microbeads. The resulting semiconductor nanoparticles were detected by fluorescence spectroscopy, microscopy, and square-wave voltammetry (SWV). The electrochemical assay based on the detection with square-wave voltammograms of Cd2+ions originating from immobilized CdS QDs showed linearity up to 45 ng mL-1, and the limit of SOD2 detection equal to 0.44 ng mL-1(1.96 × 10-11M). This detection limit is lower by 2 orders of magnitude in comparison with that of other previously published assays for superoxide dismutase. The electrochemical assay was validated with HepG2 (Human hepatocellular carcinoma) cell lysate containing SOD2.
authors.affiliation: CIC biomaGUNE , Paseo de Miramón 182, Donostia-San Sebastián 20009, Spain.
authors.forename: Ruta
authors.initials: R
authors.lastname: Grinyte
authors.affiliation: CIC biomaGUNE , Paseo de Miramón 182, Donostia-San Sebastián 20009, Spain.
authors.forename: Javier
authors.initials: J
authors.lastname: Barroso
authors.affiliation: CIC biomaGUNE , Paseo de Miramón 182, Donostia-San Sebastián 20009, Spain.
authors.forename: Marco
authors.initials: M
authors.lastname: Möller
authors.affiliation: CIC biomaGUNE , Paseo de Miramón 182, Donostia-San Sebastián 20009, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Saa
authors.affiliation: CIC biomaGUNE , Paseo de Miramón 182, Donostia-San Sebastián 20009, Spain.
authors.forename: Valeri
authors.initials: V
authors.lastname: Pavlov
mesh_terms: D000469:Alkaline Phosphatase; D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay; D056945:Hep G2 Cells; D006801:Humans; D057230:Limit of Detection; D051379:Mice; D008863:Microspheres; D045663:Quantum Dots
chemical_list: D000469:Alkaline Phosphatase
keywords: SOD2; electrochemistry; immunoassay; microbeads; quantum dots