Immuno-flow cytometric identification and enumeration of the ichthyotoxic dinoflagellate Gyrodinium aureolum Hulburt in artificially mixed algal populations

Flow cytometric identification and enumeration of Gyrodinium aureolum Hulburt (Dinophyceae) were performed in artificially mixed algal populations using direct immunofluorescence. Calibration of the flow cytometer, performed with a mixed algal population spiked with immunofluorescently labelled G. aureolum cells, showed that selection of target cells after analysis on green and orange fluorescence can be done with a recovery of 91.8% [coefficient of variation (CV) = 0.09]. Other selection methods were less good, with 67.4% (CV = 0.16) and 58.4% (CV = 0.43) recovery based on green and red fluorescence or green fluorescence and perpendicular light scattering. For mixed algal populations spiked with unlabelled G. aureolum cells, the quantification of target cells was quite good (recovery of 76.7%; CV = 0.20). The percentage of total cell loss was high (58.0-92.0%), but this was caused mostly by loss of species smaller in size than G. aureolum. Estimates of the relative contribution of G. aureolum in labelled samples were therefore often far too high, but detection and quantification were not affected. The methodological underestimation (23.3%) was partly caused by gating on green and orange fluorescence (inaccuracy 8.2%)