Magnetic Particle-Based Enzyme Assays and Immunoassays for Microcystins: From Colorimetric to Electrochemical Detection.

Abstract

In this work, magnetic particles (MPs) are used as supports for the immobilization of biorecognition molecules for the detection of microcystins (MCs). In one approach, a recombinant protein phosphatase 1 (PP1) has been conjugated to MPs via coordination chemistry, and MC-LR detection has been based on the inhibition of the enzyme activity. In the other approach, a monoclonal antibody (mAb) against MC-LR has been conjugated to protein G-coated MPs, and a direct competitive enzyme-linked immunoparticle assay (ELIPA) has been then performed. Conjugation of biomolecules to MPs has been first checked, and after optimization, MC detection has been performed. The colorimetric PPIA with PP1-MP and the best ELIPA strategy have provided limits of detection (LOD) of 7.4 and 3.9 μg/L of MC-LR, respectively. The electrochemical ELIPA has decreased the LOD to 0.4 μg/L, value below the guideline recommended by the World Health Organisation (WHO). The approaches have been applied to the analysis of a cyanobacterial culture and a natural bloom, and MC equivalent contents have been compared to those obtained by conventional assays and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Results have demonstrated the viability of the use of MPs as biomolecule immobilization supports in biotechnological tools for MCs monitoring.

Evaluation of Coupling Protocols to Bind Beta-Glucosidase on Magnetic Nanoparticles

“…[14][15][16][17][18][19][20][21][22][23][24] For example, Hernandez and coworkers 16 developed an amperometric flow system based on a magnetically immobilized glucose oxidase, for the determination of glucose in blood samples. The amino modified, silica coated, magnetite was used to bind the enzyme with the glutaraldehyde protocol.…”

“…The authors declared that their flow system gave results in accordance with an official methodology based on enzymatic spectrophotometry, 16 and the magnetic support was successfully exploited to retain the enzyme in the desired location of the amperometric apparatus. Reverte and colleagues 19 used commercial superparamagnetic particles to immobilize a recombinant protein phosphatase, exploiting the strong coordination between the histidine residuals of the recombinant enzyme and the Ni 2+ -iminodiacetic acid complex on the magnetic particles. This supramolecular complex has been tested with a microcystin-peroxidase system inhibition assay, giving good results in terms of Delivered by Ingenta to: Nanyang Technological University IP: 46.148.…”

Magnetic bead and gold nanoparticle probes based immunoassay for β-casein detection in bovine milk samples

“…After 5 min substrate incubation, 50 μL of MBs probe provided appropriate absorbance values (about 1.0). Higher amounts of the MBs probe were not tested because they would have implied the use of higher immunoreaction substance concentrations and consequently higher limit of detections (Reverte et al, 2013).…”

Rapid, High‐Resolution Magnetic Microscopy of Single Magnetic Microbeads

“…Magnetic beads are in use in biomedical and environmental sciences for a wide range of applications including enzyme immunoassays, magnetic cell separation, detection of environmental trace elements, hyperthermia, and targeted drug delivery . The different applications of magnetic beads necessitate different physical properties (i.e., size, shape, and magnetic properties) and novel applications are continually being explored.…”