Celltiter glo cell viability assay kit

Cell viability and drug sensitivity were measured using a CellTiter-Glo® Cell Viability Assay Kit (Promega) according to the manufacturer’s protocol. Briefly, cells were cultured in sextuplicate in 96-well plates (∼250 cells per well) and incubated for 24 h to allow cell attachment on the surface of the wells in charcoal-stripped serum. Then, cells were exposed to different concentration of drugs. The effects on cell viability were tested when drugs present in the culture medium. On day six, cell viability was assessed by adding 100 μL per well of cell titer glow (Promega) followed by a 10 min incubation at 37 °C and measured by the amount of luminescence using a 96-well plate luminometer (GlowMax; Promega). Background was subtracted using the medium-only control wells.

End-point cell viability assays were performed by using either Cell Titer-Glo cell viability assay kit (Promega) or WST-1 reagent (Roche, Penzberg, Germany) as manufacturer’s protocol and were measured using Synergy HT microplate reader (BioTek, Vermont, USA). Real-time cell viability, migration and invasion assays were done on xCELLigence RTCA-DP (ACEA Biosciences Inc., CA, USA) according to manufacturer’s protocol. All RTCA measurements were normalized to transfection time, and statistical significance of the results was tested by paired two-tailed student t-test. Cell cycle analysis was performed using BrdU/7AAD flow kit (BD Biosciences, USA) as previously described28 (link). For wound healing assay, 2.5 × 10⁵ cells per well were seeded in 24-well plates, and after 48 hours of incubation following the transfections and scratch formation, images were taken with 5x magnification using Nikon Eclipse inverted microscope (Nikon, Japan). All graphics and statistical analysis were carried out in GraphPad software (GraphPad software Inc., La Jolla, CA, USA).

Cells were seeded at 3 × 10⁴ cells/well in 48-well plates and allowed to proliferate until 60%–80% confluent. Complexes of phage vectors and transfection reagents prepared at optimal ratios in serum-free media, or control phage vector alone were applied to the cells, followed by incubation at 37 °C for 4 h. Next complete media, containing FBS, was administered and the cells were incubated at 37 °C to allow for transgene expression. Transduction efficiency of the phage complexes was determined by using phage carrying the green fluorescent protein (GFP) or firefly luciferase (Luc) reporter genes. GFP expression was evaluated using a Nikon eclipse TE200-S fluorescent microscope (Nikon, Surrey, UK). Luc reporter gene expression in transduced cells was determined by using the Promega Steady-Glo^® Luciferase Assay kit following the manufacturer’s protocol and quantified using a Promega plate reader (Promega, Southampton, UK). Data were normalized to 100 µg protein levels as determined by the Bradford assay and presented as relative luminescence units per 100 µg protein. Cell viability was analyzed by CellTiter-glo^® cell viability assay kit; following the manufacturer’s protocol and quantified using a Promega plate reader. All cell transduction experiments were repeated three times and performed in triplicates.