Fig. 5.

Evaluation of applicability of different fluorescent dyes for the detection of membrane permeability and membrane depolarisation in the AS mix system. Different fluorescent dyes were tested for usability to measure membrane permeability: SYTOX Green (a), TO-PRO (b); and membrane depolarisation: DIBAC₄(3) (c). Fluorescence of indicator dyes in reaction mixtures containing 50 μM AS, 50 μM H₂O₂, and 0.72 U/ml POX, and various control mixtures from which one or two components were omitted was measured. Fluorescent dyes were added either immediately (0 h) or 3 h after preparation of the mixtures to see if bleaching occurs. HK P. syringae pv. tabaci suspension was also added 3 h after mixture preparation to 5 × 10⁶ CFU/ml final density. Fluorescence was measured 4 h after preparation of the mixtures. Error bars indicate standard deviations. Asterisks indicate significant difference from corresponding controls according to student’s T-test (*p < 0.1; **p < 0.05). Abbreviations: CONT: untreated bacteria, AS: acetosyringone, HK: heat-killed, POX: horseradish peroxidase, AU: arbitrary unit