Construction strategies and secretion analysis of DENV1-4 VLPs. A) Schematic representation of three strategies to construct DENV1-4 VLP expression plasmids, pD1-D4prME include the entire prM and E gene regions of DENV. JEV signal sequence (JESS) is introduced to the N-terminal of pJD1-D4prME, and pJD1-D4prMEΔ20%JEV. The corresponding JEV-E sequence was constructed in pJD1-D4prMEΔ20%JEV to replace the 20% C-terminal region of DENV-E; B) Western blot analysis of DENV1-4 VLPs secretion, Concentrated culture supernatants from 293T cells transformed with pD1-D4prME, pJD1-D4prME or pJD1-D4prMEΔ20%JEV were tested by rabbit polyclonal serum specific against dengue E protein. Bound antibodies were detected by a HRP-conjugated goat anti-mouse antibody. The band corresponding to envelope (E) is indicated with an arrow to the right of the blot.