Melanoma Cell Line Authentication and Culture

The A375 human melanoma cell line was purchased from ATCC and cultured in DMEM complemented with 10% fetal bovine serum (FBS) (Cambrex) and 100 U/ml penicillin-streptomycin (Invitrogen). In order to authenticate the cell lines, the expected major genetic alterations were verified by NGS sequencing. The absence of Mycoplasma contamination was verified every 3 weeks with the MycoAlert detection kit (Lonza). Previously described patient-derived short-term cultures ( < 10), GLO and C-09.10, established from BRAF^V600 metastatic melanomas [21 (link)], were grown in RPMI complemented with 10% FBS and 100 U/ml penicillin-streptomycin. TNFα (100 ng/mL) and TGFβ (20 ng/mL) (Peprotech) were replaced in the culture medium every 3 days. The BRAF inhibitor PLX4032/vemurafenib was purchased from Selleck Chemicals (Houston, TX, USA) and reconstituted in DMSO.