Viruses and cells.

The murine mastocytoma cell line P815 (ATCC TIB-64) and HEp-2 cells (ATCC CCL-23) were purchased from the American Type Culture Collection and maintained in Dulbecco's modified Eagle medium (DMEM) (EuroClone, Milan, Italy) supplemented with 10% heat-inactivated fetal bovine serum (FBS) (EuroClone), 100 μg/ml streptomycin, and 100 U/ml penicillin (EuroClone) at 37°C. Vero cells (ATCC CCL-81) were maintained in DMEM supplemented with 10% FBS at 37°C. The producer cell line V3-10 (AmVac Research GmbH), a Vero cell line stably transfected with the Sendai virus P protein to transcomplement the replication-deficient Sendai virus (31), was maintained in DMEM supplemented with 10% FBS plus 200 μg/ml hygromycin B (AppliChem GmbH, Darmstadt, Germany); during vaccine production, no serum or antibiotics were added to the culture medium. BSR-T7 cells, stably transfected BHK cells that constitutively express T7 polymerase, were maintained in DMEM supplemented with 10% FBS plus 1 mg/ml G418 (AppliChem). Replication-competent Sendai virus (SeV-GFP) was produced in Vero cells at 33°C. The replication-deficient Sendai virus vaccine strain SeV76 was cultured at 33°C in V3-10 cells. All Sendai virus vectors descend from the Fushimi strain (Sendai D52). UV-inactivated vaccine SeV76 was obtained after UV irradiation at 48 mJ/cm² (254 nm) with 700 μl virus suspension in a 6-well plate. Complete inactivation was verified via viral growth and titration.

Human RSV type A (strain Long; ATCC VR-26) was cultured and amplified on HEp-2 cells at 35°C. Viral stock was titrated and stored at −80°C.