Calf serum

RS cells were a gift from E. Wagner and were propagated in minimal essential medium with Eagle's salts (MEM) (Life Technologies/Thermo Fisher Scientific) supplemented with 5% heat-inactivated calf serum (Atlanta Biologicals, Lawrenceville, GA), 292 mg/ml l-glutamine, 250 U/ml of penicillin, and 250 μg/ml of streptomycin (Life Technologies). Vero cells (procured from the American Type Culture Collection [ATCC]) and Vero cell-derived E5 cells (53 (link)) (obtained from N. DeLuca) were propagated in Dulbecco's modified Eagle's medium (DMEM) with 10% heat-inactivated fetal calf serum, 250 U/ml of penicillin, and 250 μg/ml of streptomycin. HSV-1 strain 17syn+ and ICP4 deletion strain KD6 (37 (link)) were obtained from J. Stevens. Stocks of KD6, HSV-GS3, and HSV-GS7 were propagated on E5 cells. For HSV-GS3 and HSV-GS7, the medium was supplemented with 10 nM mifepristone, and infected cultures were subjected to daily heat treatment at 43.5°C for 30 min for three successive days. EIV Prague/56 was a gift from T. Lengsfeld and was propagated on Madin-Darby canine kidney (MDCK) cells (ATCC) in DMEM supplemented with 10% heat-inactivated horse serum as previously described (54 (link)). Stocks of EIV were titrated using 50% tissue culture infective dose (TCID₅₀) assays in MDCK cells. All the cells were cultured at 37°C under 5% CO₂.

All cell lines were maintained at 37 °C under humidified conditions and 5% CO2. The N9 cells were maintained in RPMI (Corning, Corning, NY, USA) with 10% calf serum (Atlanta Biologicals, Flowery Branch, GA, USA). The HMC3 cells were maintained in DMEM medium (Corning, Corning, NY, USA) supplemented with 10% calf serum. CHO cells were maintained in DMEM:F-12 medium (Corning, Corning, NY, USA) 1:1 with 10% calf serum. SHSY5Y cells were maintained in DMEM:F12 (Corning, Corning, NY, USA) 1:1 with 10% calf serum and non essential amino acids (Thermo Fisher Scientific, Waltham, MA, USA).