Cell Colony Formation Assay

A cell colony formation assay was carried out as described in previous study [61 (link)]. In detail, stable CYP27C1-knockdown A549 and H1975 cells and the corresponding control cells were seeded in 6-well plates with a density of 2000 cells per well. Cell colonies, formed after around 14 days, were fixed by methanol at room temperature for 10 min, followed by staining with crystal violet (0.5%, w/v) of for 10 min. The images of stained cell colonies were captured by a digital scanner (V370 Photo, EPSON, Long Beach, CA, USA). The stained cell colonies were then eluted by acetic acid (10%, v/v), and the absorbance of eluent was measured by a microplate photometer at 595 nm.