A Recombinant Influenza A/H1N1 Carrying A Short Immunogenic Peptide of MERS-CoV as Bivalent Vaccine in BALB/c Mice

Shehata, Mahmoud M.; Kandeil, Ahmed; Mostafa, Ahmed; Mahmoud, Sara H.; Gomaa, Mokhtar R.; El-Shesheny, Rabeh; Webby, Richard; Kayali, Ghazi; A. Ali, Mohamed

The novel rescued virus chimeric bivalent 5 + 3 and WT 5 + 3 were propagated in 10-day-old SPF embryonated chicken eggs. MERS-CoV was grown in Vero-E6 cells. The collected harvest for each virus was clear up from cell debris by centrifugation at 800× g for 5 min. Harvested viruses were inactivated using 0.1% formaldehyde and mixed well for 24 h at 4 • C. Inactivated viral particles were pelleted by careful layering of 30 mL of inactivated virus onto 12 mL of 20% sucrose in a centrifuge tube and then centrifuged in ultracentrifuge Sorvall MTX 150 (Thermo, Waltham, MA, USA) at 28,000 rpm at 4 • C for 2 h. The pellets were suspended in 500 µL 1× PBS and virus titers were measured by HA assay. Total Protein was measured using NanoDrop 2000 (Thermo Fisher Scientific, Waltham, MA, USA). Equal protein content of each inactivated virus was mixed with alum adjuvant 1:1 (v/v) for 30 min then these suspensions were used for immunization.