Hek293t crl 3216

The following cell types, HEK293T (CRL-3216), Vero (CCL-81), Hs 729T (HTB-153), THP-1 (TIB-202) and T84 (CCL-248), were obtained from the ATCC (Manassas, VA, USA). HEK293 cells were purchased from Microbix Biosystems Inc (Mississauga, ON, Canada), and 911 cells were already described [23 (link)]. HEK293T-hACE2 cells were already described [24 (link)]. All the cell lines were grown in the recommended medium supplemented with 15% of fetal bovine serum (Natocor, Cordoba, Argentina), 2 mM glutamine, 100 U/mL penicillin and 100 μg/mL streptomycin and maintained in a 37 °C atmosphere containing 5% CO₂. For the HEK293T and HEK293T-hACE2 cell cultures, non-essential amino acids (1X final concentration) were added. Immature dendritic cells (iDC) were generated from THP-1 monocytes as previously described [25 (link)]. To induce differentiation, THP-1 monocytes were cultured during 5 days in a RPMI-1640 medium (Gibco, Whaltman, MD, USA), 2-mercaptoethanol (0.05 mM final concentration; Gibco, Whaltman, MD, USA) and fetal bovine serum (10%), adding rhIL-4 (100 ng = 1500 IU/mL; Peprotech, Cranbury, NJ, USA) and rhGM-CSF (100 ng = 1500 IU/mL; Peprotech, Cranbury, NJ, USA). The acquired properties of iDCs were analyzed under a microscope. A medium exchange was performed every 2 days with a fresh cytokine-supplemented medium.

BHK-21, HeLa (CCL-2; ATCC), HEK293T (CRL-3216; ATCC), Vero E6 cells (a kind gift from C Drosten, Charité Berlin), and TZM-bl cells (obtained from the NIH AIDS Reagent Program) were cultured in DMEM with 10% FBS, 100 µg/ml streptomycin, and 2 mM L-glutamine. For an agarose-overlay, pre-warmed DMEM with 2% FBS mixed with liquid SeaPlaque Agarose (Lonza) to a final concentration of 0.8% agarose was added to cells 2 h postinfection. For plaque assays, Vero E6 cells were overlayed with 2.5% Avicel (Merck) after 1 h of virus inoculation. HEK293T cells expressing vector or IFITM-HA proteins were generated via retroviral transduction and subsequent puromycin selection. Interferon stimulation was performed using indicated concentrations of Roferon (Interferon-α2a; Roche).

HEK293T (CRL-3216, ATCC) and HeLa (CCL-2, ATCC) cells were purchased from Shanghai Cell Bank and tested negative for mycoplasma contamination. These cell lines were authenticated using Short Tandem Repeat (STR) analysis by Shanghai Biowing Applied Biotechnology Company. HEK293T and HeLa cells were cultured in DMEM medium supplemented with 10% FBS in a 37°C humidified incubator. Cell transfections were conducted using Lipofectamine 2000 (11668019, Invitrogen) following the manufacturer's instructions.