Abstract
An amperometric immunoanalysis system based on monoclonal antibodies immobilized on Sepharose beads and packed into a micro-immunocolumn was developed for the quantification of microcystin-LR. Microcystin-LR (MCLR) was used as a reference microcystin variant. Inside the immunocolumn, free microcystins and microcystin-horseradish peroxidase (tracer) were sequentially captured by the immobilized antibodies, and the detection was performed electrochemically using Super AquaBlue ELISA substrate 2,2′-azinobis(3-ethylbenzothiazoline-sulfonic acid) (ABTS). The ABTS●+ generated by enzymatic oxidation of ABTS was electrochemically determined at a carbon working electrode by applying a reduction potential set at 0.4 V versus Ag/AgCl reference electrode. The peak current intensity was inversely proportional to the amount of analyte bound to the immunocolumn. The amperometric flow-ELISA system, which was automatically controlled through the CapSenzeTM (Lund, Sweden) computer software, enabled determination of MCLR as low as 0.01 µg/L. The assay time was very short (20 min for one assay cycle). In addition, the electrochemical signals were not significantly affected by possible interferences which could be present in the real samples. Along with the simplicity of automation, this makes the developed method a promising tool for use in water quality assessment.
Highlights
The global increase of potentially harmful cyanotoxins in surface water systems is instrumental to the search for simple, rapid and field-applicable analytical techniques for extensive freshwater monitoring [1,2].Microcystins (MCs) are a group of hepatotoxic cyanotoxins with more than 80 variants commonly present in watercourses [3,4,5,6]
Super AquaBlue was purchased from eBioscience (San Diego, CA, USA), MCLR standard was from Enzo Life Sciences (Malmö, Sweden), Adda-specific monoclonal antibodies were from Abraxis
Since strong attachment of the antibody to the support surface is required for online flow and/or sequential injection analysis, monoclonal antibodies (mAbs) were covalently immobilized on the CNBr-activated Sepharose sequential injection analysis, mAbs were covalently immobilized on the CNBr-activated Sepharose beads
Summary
The global increase of potentially harmful cyanotoxins in surface water systems is instrumental to the search for simple, rapid and field-applicable analytical techniques for extensive freshwater monitoring [1,2]. Microcystins (MCs) are a group of hepatotoxic cyanotoxins with more than 80 variants commonly present in watercourses [3,4,5,6]. MCs have a common cyclic structure built of seven amino acids including the unique amino acid called. Health risks posed by these MCs have intensified the search for convenient and rapid techniques for their detection and removal [13]. E.g., conventional enzyme-linked immunosorbent assays (ELISA) have rapidly gained commercial interest, and are presently widely used due to their robustness and reliable results. Microtitre-based ELISA kits that only target a few MC
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