Celltiter 96 aqueous one solution cell proliferation assay mts assay kit

Cells were incubated with recombinant anti-ErbB2 mAbs for 2 h, followed by the addition of ErbB ligands or not. Recombinant human EGF and HRG were added at a final concentration of 5 and 1 nM, respectively. After an additional 4-day incubation, cell proliferation was determined by CellTiter 96 AQueous One Solution Cell Proliferation Assay (MTS assay) kit (Promega, Madison, WI, USA). All measurements were performed in triplicate.

Cells were seeded at a density of 5 × 10³ cells per well in a flat-bottomed 96-well plate in humidified 37°C and 5% CO₂ atmosphere. After 12 hours, the cells were treated with 10μg/ml control IgG, trastuzumab, pertuzumab, trastuzumab plus pertuzumab, and H2-18 for an additional 5 days. Cell viability was determined by CellTiter 96 AQueous One Solution Cell Proliferation Assay (MTS assay) Kit (Promega).

To assess for GNP toxicity on GBM alone, cell viability was measured at 3 and 24 h post GNP exposure using a CellTiter 96 Aqueous One Solution cell proliferation assay kit MTS assay (Promega, Madison, WI, USA). Cells of 5 × 10³ (100 μL per well) were seeded in triplicates with 200 μL of DMEM in the 96-well plates and stored in the incubator overnight in a humidified atmosphere at 37 °C with 5% CO₂. Next, the cells were washed with PBS and the medium was replaced with fresh medium containing different concentrations of GNPs (50, 100 μg/mL) and medium only (GBM cell with no GNP). After 3 h, the medium was removed and cells were washed with PBS twice. Next, 20 μL MTS was added to each well and incubated for 1 h. The optical density (OD) was recorded at 490 nm in a 96-well plate reader (Biorad, Watford, UK). The experiment was repeated to assess for cytotoxicity at 24 h. Cell viability was expressed as a percentage of the absorbance value of the GNP treated group to the GBM alone group. Data was a representation of 3 independent experiments (n = 3) for each group (3 and 24 h post GNP exposure).