Crystal violet solution

The long-term proliferation of cells could be detected by their ability to form colonies. Firstly, seeded 400 MM cells per 6-well plate. After 14 days, we removed the cell culture medium and gently washed the cell culture plate containing the colonies with PBS, then fixed them for 20 min using 3.7% formaldehyde (Meilunbio, China). The colonies were washed with PBS again, stained with crystal violet solution (Meilunbio, China) for 10 min and their numbers were measured (relative clonality = number of clones/average number of clones in the control group).

RPMI8226-oeCON, RPMI8226-oeNCX1, KMS11-oeCON and KMS11-oeNCX1 cells were seeded in 6-well plates (400 cells/well) and fed with RPMI 1640 complete medium in the presence or absence of chloroquine (CQ) or BTZ (Supplier name: Velcade) 3 times a week. After 14 days of observation, one colony was defined as more than 40 cells. Cells were then fixed with 3.7% formaldehyde (Supplier name: Meilunbio) for 20 min, followed by staining with crystal violet solution (Supplier name: Meilunbio;Catalogue number:MB4721) for 10 min. The plate was imaged and colonies were counted. Each sample was repeated three times.