FIG 2.

Diagram of the WEEV genome, E1ecto construct, and LANAC formulation. (A) The WEEV genome contains a 5′ untranslated region (5′UTR), nonstructural polyprotein genes (nsp1-nsp2-nsp3-nsp4), subgenomic promoter sequence, structural polyprotein genes (capsid-E3-E2-6K-E1), and 3′ untranslated region (3′UTR). (B) The E1ecto construct encoded full-length WEEV (McMillan) 6K, the first 408 amino acids of WEEV (McMillan) E1, a tobacco etch virus protease cleavage site (TEV), and an 8× histidine purification tag. E1ecto was produced using the baculovirus-insect cell system. (C) E1ecto or a sham preparation (see Materials and Methods) was mixed with PIC- and ODN-containing liposomes to form E1ecto or sham LANACs that were used for vaccination experiments. (D) Optimal E1ecto antigen dose in LANAC. Mice (n = 10/group) were vaccinated with LANACs containing 0.1, 1.0, or 10 μg of E1ecto using a prime-boost protocol and then challenged by s.c. inoculation with 10⁴ PFU of WEEV Montana-64. Complete protection was observed using the 10-μg antigen dose, and this dosage was used for the remainder of the vaccination experiments.