High extracellular iron concentrations increased intracellular ROS levels. (A) Determination of intracellular ROS production. WT cells were exposed to 0 (H2O control), 1 or 30 μM FeCl3 in RPMI at 30°C for 10 min. Additionally, cells were exposed to 30 μM FeCl3 together with 10 mM NAC. Means and standard deviations are shown from one representative experiment where all samples were derived from the same pre-culture. ** denotes P ≤ 0.01 (student’s t-test). All experiments were repeated 2 – 4 times from independent pre-cultures with similar results. (B) Influence of ROS on flocculation. Flocculation of cells was triggered by 30 μM FeCl3 in RPMI with or without 10 mM NAC. After 2 h incubation at 30°C, sedimentation rates were determined as described in the experimental part. Means and standard deviations of three independent samples are shown (n = 3).