Streptomycin penicillin amphotericin solution

Porcine Reproductive and Respiratory Syndrome virus type 1 (PRRSV-1) strain PRRS-FR-2005-29-24-1 (Finistère strain; genotype 1.1) was propagated on AM cultured in RPMI 1640 medium supplemented with 10% FCS and 1% Streptomycin/Penicillin/Amphotericin (SPA) solution (Eurobio scientific, Les Ulis, France) for 72 h. The supernatant was clarified by centrifugation for 20 min at 600× g and then purified on Amicon Ultra-15 centrifugal Filters (Sigma-Aldrich, Saint-Quentin Fallavier, France) after a 20 min centrifugation at 4000× g and 4 °C. The PRRSV-1 titer using Tissue Culture Infectious Dose (TCID50) assay protocol on AM was 8.7 × 10⁶ TCID50/mL. The Aujeszky’s disease virus (ADV) strain Kojnok propagation was p45to5m4r on Newborn Pig Trachea (NPTr) cells [25 (link)] in DMEM medium supplemented with 10% FCS and 1% of SPA solution (Eurobio scientific). The ADV titer using TCID50 assay protocol on Madin-Darby Canine Kidney cell line (MDCK) was 3.7 × 10⁷ TCID50/mL.

NPTr cells [17 (link)] were cultured with Dulbecco’s modified Eagle medium (DMEM) (Eurobio scientific, Les Ulis, France) supplemented with 10% fetal calf serum (FCS) (Eurobio scientific) and 1% of Streptomycin/Penicillin/Amphotericin (SPA) solution (Eurobio scientific). Flasks were then incubated at 37 °C in a 5% CO₂ humidified environment and sub-passages were carried out once a week to assure the cell line maintenance.