E plate 96 pet

Growth curves of embryonal CNS tumor cells were determined by the impedance‐based xCELLigence real‐time cell analysis system (ACEA Biosciences, San Diego, CA, USA). Briefly, 50 µL of cell culture media was added to each 96 well of the E‐Plate 96 PET (ACEA Biosciences) for background reading. Subsequently, 50 µL of cell suspension containing 2000 cells was added to each well and the plate was placed on xCELLigence station inside the incubator. Twenty‐four hours later, cells were subjected to miR‐367 silencing and impedance reflecting cell adhesion and proliferation changes was measured every 15 min for 7 days. Data are expressed as changes of impedance (‘Cell Index’) over time, according to the manufacturer’s instruction.

A non-invasive, impedimetric technique was utilized to validate the results of the endpoint cell viability assay in A2058 cells. The adherent cells were seeded (10⁵ cells/mL) in a special 96-well plate (E-Plate 96 PET; ACEA Biosciences, San Diego, CA, USA). The bottom of the wells is covered by gold microelectrodes that can register the changes in impedance in an electrically conductive solution. These changes are proportional to the number of cells; thus, the antiproliferative effects of the compounds can be measured. Before seeding the cells, a baseline with a cell-free culture medium was assessed. Following overnight incubation, the cells were treated with 13.5 nM BOZ + 13.5 μM TIC10, 13.5 nM BOZ + 40.5 μM TIC10 and the matching single agents and controls (medium and DMSO). The data were registered by the RTCA 2.0 software (Real Time Cell Analyzer; ACEA Biosciences, San Diego, CA, USA). This software is capable of converting the measured impedance data to a unitless cell index (cell index = (Z_ti − Z_t0)/F_i; Z_ti: impedance at individual time point; Z_t0: impedance at start of experiment; F_i: constant). The experiment was performed in triplicates, and the results were normalized to the untreated medium control and represented as mean ± SD.