Fig. 4.
C2IN-C3lim inhibits chemotaxis of primary human monocytes ex vivo. Primary human monocytes were isolated from healthy donors and 1.6 × 105 cells per sample were incubated at 37 °C for 1 h in RPMI medium without FCS with or without the following toxins: C2I (1 µg/mL) + C2IIa (2 µg/mL), C2IN-C3lim (1 µg/mL) + C2IIa (2 µg/mL), C2IN-C3lim (1 µg/mL) and C2IN-C3lim (2 µg/mL). During the last 20 min of incubation, the fluorescence dye BCECF was added into the medium to label the cells. Subsequently, cells were applied into the upper wells of the trans-well Boyden chamber and incubated for 40 min towards the chemoattractant fMLP (100 nM) in the lower chamber wells. For control, cells were not exposed to fMLP. The membrane filters between the upper and lower chambers had a pore size of 8 µM. Chemotaxis was monitored by measuring the fluorescence signal of the cells associated with the filter after a 40 min incubation period. Values are given as mean ± SD (n = 6) and significance was tested for each sample against the “fMLP-treated control group” using Student’s t test (*p < 0.05, **p < 0.01, ***p < 0.001)