Dmem glutamax medium

In brief, HEK-293 cells were grown in Petri dishes in Dulbecco’s modified Eagle’s medium (Gibco Thermo Fisher, Waltham, MA, USA) supplemented with 10% FCS (Biochrom GmbH, Berlin Germany). Cells were routinely passaged twice a week and incubated at 37°C under 5% CO₂ growth conditions. tsA-201 cells were cultured at 37°C and 5% CO₂ in a 60-mm-diameter cell culture dish in 4-ml DMEM GlutaMAX medium (Biochrom GmbH, Berlin Germany) with 10% FCS (Biochrom GmbH, Berlin Germany) and penicillin-streptomycin (Sigma-Aldrich, St. Louis, MO, USA). Cells were routinely passaged twice a week. HEK-293 and tsA-201 cells were transfected using standard calcium phosphate method (Koch et al. 2016 (link)). For whole-cell and single-channel recordings, HEK-293 cells were transfected with a 1:0.5:1.5 ratio of Ca_V1.2 (α1c#77, NM_001129843.2) or Ca_V2.2 (NM_000718.3), either human WT (NM_000723) or mutant Ca_Vβ-subunit and human Ca_Vα₂-δ₁-subunit (NM_000722.3). For co-immunoprecipitation experiments, tsA-201 cells were transfected with a 1:2 ratio of either human WT or mutant Ca_Vβ_1b_HA-subunit and human Flag-Gem.