CBP Release Package — page 5

PEDv Virus Expansion October 2014 a: PEDv can be grown on Vero-76 or Vero-81 cell lines. The virus grows at the same efficiency in 2) conditions. Vero-81 cells seem to be more resilient to Trypsin and may be the preferred cell line for assays requiring long incubations in the PEDv Maintenance Media. Materials: ooo00o00o000 eooo000000 Safety: PEDv Maintenance Media 5m! Sterile Individually wrapped pipettes for Pipet Aid 10ml Sterile Individually wrapped pipettes for Pipet Aid Sterile 1x PBS (no Ca, Mg) 70% EtOH Water bath set at 37°C 500ml Plastic waste container Gloves Laminar flow hood Paper Towels Confluent flasks of Vero cells 1mL of PEDv Conical holder/Rack Sterile 50ml conicals Microcentrifuge tubes Wear gloves and lab coat while performing this procedure. This procedure is performed in a laminar flow hood Complete all training required by the University of Minnesota and the Department of Environmental and Health & Safety guidelines Method 35) 2) 3) 4) Heat PBS in water bath for 15 minutes Bring PEDv aliquot to room temperature Sterilize workspace in laminar flow hood by spraying area with 70% EtOH and wiping with paper towels Sterilize Pipet Aid and plastic waste container by spraying with 70% EtOH and wiping with paper towels prior to placing in hood.