Generation of monoclonal antibodies for the specific immunodetection of the toxic dinoflagellate Alexandrium minutum Halim from Spanish waters

In recent years, Alexandrium minutum (A. minutum) has provoked repeated toxic episodes of harmful algal blooms. The existing method for accurate identification of this dinoflagellate is based on morphological traits and requires taxonomic expertise. However, this is impractical when rapid identification of toxic strains is required, when the microalgae are in low density or when automatic monitoring is needed. In this study, the generation and application of two monoclonal antibodies (M90.3 and M54.3) directed against an intracellular antigen of the A. minutum strain AL1V are described. Specificity and cross-reactivity were tested with cells from clonal cultures. In an immunofluorescence assay, confocal microscopy and flow cytometry showed that the antibodies specifically labeled the two tested A. minutum strains (AL1V and AMP13) and did not show cross-reaction with other Alexandrium species, such as Alexandrium tamarense SZN19, A. tamarense MDQ1096, Alexandrium catenella IEO715 and Alexandrium andersonii SZN12, or tested members of other phylogenetic classes. Furthermore, M90.3 showed highly specific recognition when used for the detection of A. minutum in natural seawater samples. Thus, the mAbs presented here could be used in microalgae monitoring, allowing a specific unequivocal identification of the toxic dinoflagellate A. minutum.