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 Cd ions originating from immobilized CdS QDs showed linearity up to 45 ng mL, and the limit of SOD2 detection equal to 0.44 ng mL (1.96 × 10 M). 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.

Preparation of Highly Stable and Photoluminescent Cadmium‐Free InP/GaP/ZnS Core/Shell Quantum Dots and Application to Quantitative Immunoassay

“…Indium phosphide (InP) quantum dots (QDs), as one of nontoxic, eco-friendly and cadmium (Cd)-free colloidal QDs, have been attracting continuous attention recently, because of the consideration as an ideal alternative material to the most www.advancedsciencenews.com www.particle-journal.com introduction in the field of IVD, optical biosensors based on fluorescent QD-based materials have achieved significant developments. [25][26][27][28] However, to be used in biomedical fields, QDs should be excellently biocompatible and maintain strong and stable fluorescence under complex physiological environments. [29,30] Theoretically, the growth of a thick shell over core is needed to passivate surface effectively and enhance the fluorescence stability.…”

“…Among various QD‐based applications, biomedicine is one of the fast developed field, including cellular labeling, gene sequencing, in vivo imaging, and in vitro diagnostic (IVD), etc. In particular, with the QDs fluorescent labels introduction in the field of IVD, optical biosensors based on fluorescent QD‐based materials have achieved significant developments . However, to be used in biomedical fields, QDs should be excellently biocompatible and maintain strong and stable fluorescence under complex physiological environments .…”

Fractal Silver Dendrites as 3D SERS Platform for Highly Sensitive Detection of Biomolecules in Hydration Conditions

“…In the last few decades, the task of analytic detection has been extensively studied for different application fields, especially for biomolecules and specific markers, towards the analysis of dangerous analytes [1]. Most commercial sensors rely on the recognition of the target through its fluorescent labeling by using dyes [2] or quantum dots [3]. Despite their diffusion due to the high sensitivity, the use of fluorescent tags introduces complex issues related to their stability due to aggregation, chemical treatment and photobleaching, resulting in a poor reliability of the sensors [4].…”

Methods of Encapsulation of Biomacromolecules and Living Cells. Prospects of Using Metal–Organic Frameworks

“…Polyacrylamide gel is used quite often [103] despite its toxicity. Other carriers such as PVA-based cryogels [104,105], ENT/ENTP photocrosslinked polymers (polymer mixture based on polyethylene or polypropylene glycol, hydroxyethyl acrylate, and isophorone diisocyanate, which is polymerized under near UV irradiation) or modified PVA [106], granules and films based on polyvinyl chloride [107], hydrogels based on chitosan [108][109][110], polycarbamoyl sulfonate [111,112], and polyurethane [113,114], peptide polymers [115], biotin-avidin linkers [116][117][118], solgel matrices based on aluminum oxide [119] or composite polymers [120][121][122][123], as well as electropolymeriz able polyaniline films [124][125][126][127] and other compounds [128,129], and nanostructured materials (including carbon nanotubes and metal nanoparticles) [130][131][132][133][134][135][136][137] should also be noted. A common disadvantage of encapsulation in gels and polymers is diffusion restrictions imposed by the carrier nature, which in some cases appreciably reduce the rate of biocatalysis and the activity of biocomponents.…”