Rpmi 1640 antibiotic free medium

The human embryonic kidney 293 (HEK 293) cell lines (from Emmanuel Bourinet, Montpellier, France) stably expressing Ca_V3.2 or Ca_V3.3 [41 (link)] were cultured under 5% carbon dioxide at 37 °C in Dulbecco’s Modified Eagle Medium (DMEM) Glutamax (Gibco, Life Technologies, Carlsbad, CA, USA) supplemented with 10% (v/v) fetal bovine serum (FBS), 100 U/mL penicillin, 100 μg/mL streptomycin (Gibco, Life Technologies), and 750 μg/mL geneticin (G418) (Gibco, Life Technologies). The Chinese Hamster Ovary (CHO) cell lines (Emmanuel Bourinet, Montpellier, France) expressing Ca_V3.1 [42 (link)] were cultured under 5% carbon dioxide at 37 °C in Alpha Minimum Essential Media (MEM) Glutamax (Gibco, Life Technologies), supplemented with 10% (v/v) fetal bovine serum (FBS) and 300 μg/mL geneticin (G418) (Gibco, Life Technologies). The human neuroblastoma SH-SY5Y cells (Victor Diaz, Goettingen, Germany) were cultured under 5% carbon dioxide at 37 °C in RPMI 1640 antibiotic-free medium (Invitrogen, Carlsbad, CA, USA), supplemented with 15% FBS and 2 mM GlutaMAX™ (Invitrogen). D-PBS (Gibco, Life Technologies) was used to wash the cells, and 0.25% Trypsin-EDTA (Gibco, Life Technologies) was used to detach the cells from the flask surface. They were split in a ratio of 1:5 (ideally 1000 cells/cm²) when they reached 70–80% confluence (every 2–3 days).

In the present study, four prostate cancer cell lines were used. The androgen-sensitive LNCaP (DSMZ GmbH, Braunschweig, Germany) and VCaP (ATCC, Manassas, VA, USA) cells are representative of prostate adenocarcinoma and the androgen-insensitive PC-3 (ATCC, Manassas, VA, USA) and CTM-LNCaP (subline of LNCaP) cells. To establish CTM-LNCaP, the parental LNCaP cells were cultivated for at least 3 months in phenol red-free RPMI medium supplemented with hormone-free charcoal-treated FBS.
All cells were cultured in RPMI-1640 antibiotic-free medium (Invitrogen, Waltham, MA, USA), supplemented with L-Glutamine, and grown in a 5% CO₂ atmosphere at 37 °C. PC-3 and VCaP cells were cultured with 10% fetal bovine serum (FBS), CTM-LNCaP with 10% charcoal-treated fetal bovine serum (FBS), and LNCaP cells with 20% FBS.