2.4 Western blot analysis
Cells (HEK293T) were transfected with the two-plasmid system and
incubated at 37 °C until the pseudotyped EBOV was harvested. The cells
were harvested and lysed. After centrifugation, the lysates were mixed
with 4×LDS sample buffer (ThermoFisher Scientific) and denaturing at
90℃for 10 min. Samples were loaded onto to a 12% SDS-PAGE gel. After
electrophoresis, proteins were transferred onto nitrocellulose (NC)
membranes. The NC membranes were then blocked using SuperBlocking Buffer
(Thermo Fisher, TX, USA) at room temperature for 2 h and subsequently
incubated with anti-p24 antibody (Sino Biological, China) and anti-EBOV
GP antibody (rabbit serum immunized with purified, truncated EBOV GP inEscherichia coli ) for 1 h at room temperature. After three washes
with phosphate-buffered saline containing tween 20 (PBST), the NC
membranes were incubated with HRP-conjugated goat anti-rabbit IgG (Bios,
China) at room temperature for 1 h. The signals were visualized on a
Fujiflm LAS-4000 image reader (Fujiflm, Tokyo, Japan) with SuperSignal
West Dura Extended Duration Substrate Kit (Thermo Fisher).