Protocol detail

Quantitative Cell Colony Formation Assay

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Cells were plated at 250 (SK-N-BE) or 100 (U2OS) cells/well in 6-well plates, in full growth medium. Media were replaced by fresh culture medium every 4 days. After 15 days, cells were washed with PBS, fixed with methanol and stained with 0.5% crystal violet. Digital images of cell colonies were obtained using a scanning device (Epson Perfection V700 Photo). Quantification of the clonogenic assay was performed with the ImageJ plugin “ColonyArea” [27 (link)] to determine the area covered by colonies in the well and the colony intensity percentage.

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Zonta F., Borgo C., Quezada Meza C.P., Masgras I., Rasola A., Salvi M., Pinna L.A, & Ruzzene M. (2021). Contribution of the CK2 Catalytic Isoforms α and α’ to the Glycolytic Phenotype of Tumor Cells. Cells, 10(1), 181.

Publication 2021

Perfection V700 Photo

Assay Cell Crystal violet Culture medium Device Methanol

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Variable analysis

independent variables

Cell line (SK-N-BE or U2OS)

dependent variables

Area covered by colonies in the well
Colony intensity percentage

control variables

Initial cell plating density (250 cells/well for SK-N-BE, 100 cells/well for U2OS)
Culture medium (full growth medium)
Medium replacement (every 4 days)
Incubation duration (15 days)
Staining method (PBS wash, methanol fixation, 0.5% crystal violet staining)
Imaging method (digital images using Epson Perfection V700 Photo scanner)
Quantification method (ImageJ plugin 'ColonyArea')

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