Modified vaccinia Ankara vaccine expressing Marburg virus-like particles protects guinea pigs from lethal Marburg virus infection

摘要

a Advantages of the MVA-based VLP vaccine platform compared with MVA vector-based vaccines and VLP vaccines. b Vector Map. MARV gp gene was inserted between the I8R and G1L sequences of MVA, and vp40 was inserted into the restructured and modified deletion III. These insertion sites have been identified as supporting high expression and insert stability. Positions are given in kilobase pairs (kbp) in the MVA genome. c Western blot for MARV GP and VP40 expression. DF1 cells infected with a MOI of 0.5 FFU/cell of parental MVA, MVA–mVP40, or MVA–MARV–VLP. Supernatants and cell lysates were run on a 4–12% SDS-PAGE before transfer and detection with antibodies specific for MARV GP and MARV VP40, respectively. The two fragments shown are parts of the same gel. d Electron microscopic analysis of expression of virus-like particles from MVA–MARV–VLP. HEK293 cells were infected with MVA–MARV for 24 h, stained with a human anti-GP antibody, fixed with 1% glutaraldehyde in 0.1 M phosphate buffer and incubated in 50 mM glycine to block residual aldehyde. Following incubation in goat anti-human secondary antibody conjugated to ultra-small gold particles, silver enhancement was performed to increase the size of gold particles for subsequent viewing on a JEOL JEM-1400. Localization of some gold beads is indicated by black arrows.