This caused mature kleptocnides to appear out of the intensity range (blue, the colour for overload of measuring range) with an intensity value of 255 i.u. or higher. The highest fluorescence intensity of every single nematocyst within the ten sections of a cnidosac was determined (maximum absolute fluorescence of single nematocysts, fNc). In addition the background fluorescence (fBg) in every section was measured to finally calculate a relative value (fNc/fBg). This value was determined by dividing the maximum absolute fluorescence of a nematocyst by the fluorescence of the surrounding tissue of the same section (Fig. 3A). Additionally, the percentage of kleptocnides

with a higher fluorescence than 255 i.u. was calculated for every time slot (Table 1). Statistical analysis was performed Volasertib supplier with Microsoft Excel and SPSS 15 (IBM). Statistical tests on significance of the changes in fluorescence intensity values GSK1120212 over time were performed with the non-parametric Mann Whitney U test (SPSS15). The autofluorescence of unstained nematocysts in Aiptasia spec. and in Aeolidiella stephanieae was very low and negligible. The nematocysts became slightly visible when excitation was amplified with settings of PMT1 = 900 V, almost twice as much as used in the Ageladine A staining experiments. Preliminary

staining experiments revealed that Ageladine A easily permeates the nematocysts of gastropods and their only food (Fig. 1B–D, Fig. 2A, B). Undischarged nematocysts with high fluorescence intensity were present in high amounts within the Aiptasia epidermis at the tip of the tentacles and especially in the acontia ( Fig. 2A, B). Lower amounts of fluorescing nematocysts were found along the tentacles and throughout the scapus. Nematocysts exhibiting hardly any fluorescence were found in lower numbers along the tentacles, and in higher numbers in the scapus. Gastropod feeding was used to trigger the discharge of the anemone’s nematocysts. Discharged nematocyst capsules, which could be found around

the two animals (anemone and gastropod) in high numbers, lost their high fluorescence with time, although their threads continued glowing, especially when PMT1 = 500 V is chosen (Fig. 2E). This was also observed in discharged kleptocnides extruded from the cnidosac (Fig. 1D). In the digestive gland and stomach area of the freshly fed gastropod, non-fluorescing nematocysts outnumbered fluorescing ones by far. This became obvious when transmission pictures were compared to fluorescence pictures (Fig. 2C, D). Since the body with the overlying viscera in the gastropod was very thick and optic measurements were difficult, no statistic values could be obtained. To investigate properties of incorporated nematocysts in the gastropod at various times, 47 cnidosacs with 1770 nematocysts in total were measured at five intervals (7, 24, 48, 72 and 96 h) after food uptake.