Congener-Independent Immunoassay for Microcystins and Nodularins

• Published in: Environmental science & technology Vol. 35; no. 24; pp. 4849 - 4856
• Main Authors: Fischer, Werner J, Garthwaite, Ian, Miles, Christopher O, Ross, Kathryn M, Aggen, James B, Chamberlin, A. Richard, Towers, Neale R, Dietrich, Daniel R
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 15-12-2001
• Subjects: Algae; Applied sciences; Bacteria; Contamination; Continental surface waters; Cyanobacteria - chemistry; Drinking water and swimming-pool water. Desalination; Earth sciences; Earth, ocean, space; Engineering and environment geology. Geothermics; Environmental impact; Exact sciences and technology; Immunoassay; Microcystins; Natural water pollution; Peptides, Cyclic - analysis; Poisons; Pollution; Pollution, environment geology; Water Pollutants - analysis; Water pollution; Water Pollution - analysis; Water treatment and pollution; Trace analysis; Drinking water; Chemical analysis; Antibody; Cyclic peptides; Ultratrace compound; Heptapeptide; Enzyme immunoassay; Hapten; Toxin; Microbial origin; Detection limit; Cyanobacteria; Surface water; Water quality; Bacteria; Water pollution; ELISA assay; Pentapeptide
• ISSN: 0013-936X; 1520-5851
• DOI: 10.1021/es011182f

Cyanobacteria (blue-green algae) (e.g., Microcystis and Nodularia spp.) capable of producing toxic peptides are found in fresh and brackish water worldwide. These toxins include the microcystin (MC) heptapeptides (>60 congeners) and the nodularin pentapeptides (ca. 5 congeners). Cyanobacterial cyclic peptide toxins are harmful to man, other mammals, birds, and fish. Acute exposure to high concentrations of these toxins causes liver damage, while subchronic or chronic exposure may promote liver tumor formation. The detection of cyclic peptide cyanobacterial toxins in surface and drinking waters has been hampered by the low limits of detection required and that the present routine detection is restricted to a few of the congeners. The unusual β-amino acid ADDA (4E,6E-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid) is present in most (>80%) of the known toxic penta- and heptapeptide toxin congeners. Here, we report the synthesis of two ADDA-haptens, the raising of antibodies to ADDA, and the development of a competitive indirect ELISA for the detection of microcystins and nodularins utilizing these antibodies. The assay has a limit of quantitation of 0.02−0.07 ng/mL (depending on which congeners are present), lower than the WHO-proposed guideline (1 ng/mL) for drinking water, irrespective of the sample matrix (raw water, drinking water, or pure toxin in PBS). This new ELISA is robust, can be performed without sample preconcentration, detects toxins in freshwater samples at lower concentrations than does the protein phosphatase inhibition assay, and shows very good cross-reactivity with all cyanobacterial cyclic peptide toxin congeners tested to date (MC-LR, -RR, -YR, -LW, -LF, 3-desmethyl-MC-LR, 3-desmethyl-MC-RR, and nodularin).