CHO-K1 cells obtained from the American Type Culture Collection (CCL-61) and TRvb1 CHO cell line lacking the endogenous transferrin receptor and stably expressing the human transferrin receptor (48) were maintained in high-glucose DMEM/F12. HeLa cells were maintained in high-glucose DMEM. Media for cell lines were supplemented with 10% heat-inactivated FBS (Invitrogen) and 5% (v/v) of a stock solution containing penicillin (5000 U/ml) and streptomycin (5 g/ml) (Invitrogen) and cultured at 37°C and 5% CO2. For transient transfections, cells were grown to 50 to 70% confluency and transfected using Lipofectamine 2000 (Invitrogen). Generation of YFP-HA-Orai1 stable CHO cell lines was by pDS-YFP-HA-Orai1 DNA transfection and selected using selective media [DMEM/F12, 10% FBS, G418 (800 μg/ml), and penicillin and streptomycin (100 μg/ml)].

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