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Colony-forming assay (stained with crystal violet)
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Cells were washed twice in ice-cold PBS and fixed with ice-cold methanol for 10 min for permeabilization of cells. Expanded colonies were stained by 0.1% crystal violet (Sigma) and 10% ethanol in PBS solution for 5 min and PBS washed 4 times, and plate images were acquired. For quantification, the colonies were incubated with elution buffer (50% ethanol, 40% distilled water, 10% acetic acid) for 5 min and transferred to a 96-well plate; the OD at 580 nm was determined using a microplate reader.
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