Establishment and Maintenance of B16-F10 and HEK293 Cell Lines

Mouse melanoma cell line B16-F10 (CRL-6475) was obtained from American Type Culture Collection (ATCC; Manassas, VA, USA). HEK293 cells stably expressing human LAT1 and human LAT2 (designated as HEK293-hLAT1 and HEK293-hLAT2, respectively) were established as described previously through transfection with plasmids encoding human LAT1 and human LAT2^{26 (link)}. B16-F10 cells were cultured in RPMI-1640 medium (Nacalai Tesque, Kyoto, Japan)^{48 (link)}. The HEK293-hLAT1 and HEK293-hLAT2 cells were maintained in MEM medium (Nacalai Tesque, Kyoto, Japan) supplemented with 1% non-essential amino acids (Wako, Osaka, Japan) and G418 disulfate (0.9 g/L, Nacalai Tesque, Kyoto, Japan)^{26 (link)}. Both cell media were supplemented with 10% fetal bovine serum (FBS, Nichirei Biosciences Inc., Tokyo, Japan) and a combination antibiotic consisting of 100 units/mL penicillin and 100 μg/mL streptomycin (Nacalai Tesque, Kyoto, Japan). Cells were incubated at 37 °C in a humidified atmosphere containing 5% CO₂.