Cationic amphiphilic drugs are often associated with the induction of phospholipidosis, which is believed to be a precursor event to liver toxicity. When studying phospholipidosis/steatosis in vitro, IONTOX uses HepG2 cells in combination with LipidTox Red, as our primary model to discriminate between phospholipidosis (phospholipids accumulated in multilaminar bodies) and steatosis (general fat accumulation). A combination of fluorescence probes, fluorescent microscopy and fluorescent signal measured in a 96-well fluorescent reader allows this to be done.
The phospholipidiosis and steatosis assay is performed in a 96-well plate, with a 7-point dose response curve, at a single time point. Following exposure to the test material, phospholipidosis and steatosis is determined using activation of fluorescent probes, the intensity of which can be read in a plate reader at 540 nm excitation and 680 nm emission.
When run with IONTOX, the phospholipidiosis and steatosis assay includes a determination of both phospholipidosis and steatosis, positive and negative controls in HepG2 cells, at a 7 concentrations in triplicate and a single time point.