2.8. Cell Proliferation Assay

The proliferation of A549 cells was determined by the fluorescent resazurin (Sigma-Aldrich) assay as described previously [28]. Briefly, an aliquot of all types of cells (6000) pre-cultured in either different 3D or 2D conditions were removed and seeded into 96-well plates (Corning Life Sciences) in DMEM/F12 10 % FBS (Gibco) in order to perform the viability assay every day. Resazurin reagent (Sigma-Aldrich) was dissolved in PBS (pH 7.4) at 0.15 mg/mL concentration, 0.22 µm filtered and aliquoted at −20 °C. We applied resazurin 20 µL stock to 100 µL culture. After 2 h incubation at 37 °C under 5 % CO₂ (Sanyo) fluorescence (530 nm excitation / 580nm emission) was recorded on a multimode microplate reader (Cytofluor4000, PerSeptive Biosytems, Framingham, MA, USA). Proliferation was calculated with relation to blank wells containing media without cells. (Significance was compared to 2D TC, pairwise. RFU = relative fluorescence unit)