The influenza A virus strains A/Singapore/INFIMH-16-0019/2016 IVR-186 (H3N2), A/Switzerland/9715293/2013 (H3N2) (mouse-adapted), A/Philippines/2/1982 (H3N2, X-79, A/PR/8/1934 reassortant), A/duck/Czechoslovakia/1956 (H4N6, A/PR/8/1934 reassortant) (52 (link)), A/Shanghai/1/2013 (H7N9, A/PR/8/1934 reassortant), A/canine/Illinois/41915/2015 (H3N2) and A/Netherlands/602/09 (H1N1) were grown in 10-d-old embryonated chicken eggs (Charles River) for 48 h at 37 °C. After virus growth, the eggs were chilled at 4 °C overnight. The following day the allantoic fluid was harvested from the chilled eggs and cellular debris was removed via centrifugation (4,000 × g for 10 min at 4 °C). Viruses were aliquoted and stored at −80 °C. Virus titers (plaque forming units/mL) were determined via plaque assay.
A/Singapore/INFIMH-16-0019/2016 IVR-186 (H3N2), A/Philippines/2/1982 (H3N2, X-79), A/duck/Czechoslovakia/1956 (H4N6) and A/Shanghai/1/2013 (H7N9) are reassortant viruses and contain the internal genome segments of A/Puerto Rico/8/1934 (H1N1).