Novel monoclonal antibodies against microcystin and their protective activity for hepatotoxicity

Abstract

Six monoclonal antibodies (MAbs) to microcystin-LR (MCLR), a cyclic heptapeptide hepatotoxin isolated from the cyanobacterium Microcystis aeruginosa, were produced. They showed the protective effects on hepatotoxicity of MCLR in vitro and in vivo, and on the inhibition of protein phosphatase by MCLR. Competitive enzyme-linked immunosorbent assays with various microcystins revealed that the six MAbs recognized a part of the molecule, in particular, a tertial structure around Adda, 3-amino-9-methoxy-2,6,8,-trimethyl-10-phenyldeca-4,6-dienoic acid. The specificity of these MAbs varied slightly. In primary rat hepatocyte cultures, all MAbs showed protective effects against the MCLR-induced cell damages, assessed by morphological changes, lactate dehydrogenase release into the medium, and a calorimetric assay to measure the cell viability using a tetrazolium dye. The M8H5 MAb showing the highest affinity for MCLR blocked the lethal effects and hepatocellular damage to mice. In addition, M8H5 MAb recovered protein phosphatase 2A inhibition by MCLR in a dose-dependent manner, while phosphatase inhibition by okadaic acid was not affected. Thus, the MAbs specifically reacted with the microcystins and prevented their biological activities. This is the first report on the protective effects of specific monoclonal antibodies on MCLR-induced toxicity.