Cy3 goat anti rabbit immunoglobulin igg

293T and Marc-145 cells were seeded on coverslips and transfected with plasmids pEGFP-C1 and GFP-nsp2, respectively. Marc-145 cells were infected with the HP-PRRSV TA-12 strain at 0.01 MOI. At 24 h post-transfection or post-infection, the cells were fixed with 4% formaldehyde and permeabilized with 0.1% (v/v) Triton X-100 in PBS. The transfected 293T and Marc-145 cells were probed with anti-14-3-3β (Abcam, Cambridge, UK) and anti-14-3-3γ/ε/ζ (Santa Cruz Biotechnology, Dallas, TX, USA) antibodies. The 14-3-3 proteins were visualized using Cy3-goat anti-rabbit immunoglobulin (IgG; Jackson, West Grove, PA, USA).
The HP-PRRSV-infected Marc-145 cells were incubated with anti-NSP2 polyclonal antibodies and visualized using fluorescein isothiocyanate (FITC) goat anti-rabbit IgG. The anti-NSP2 antibodies were prepared by immunizing New Zealand white rabbits with a peptide composed of the N-terminal 180 amino acids of NSP2 (NSP2-180), which had been produced previously [27 (link)]. The activity of these antibodies was confirmed by Western blot and immunofluorescent and enzyme-linked immunosorbent assays (data not shown). All probed cells were observed under a fluorescence microscope (Leica, SPE, Buffalo Grove, IL, USA).

DEF cells were seeded on coverslips and co-transfected with pEGFP-LC3 and 10 recombinant plasmids as well as empty vectors, respectively. At 24 hpt, the cells were fixed with 4% formaldehyde and permeabilized with 0.1% (v/v) Triton X-100 in PBS. The transfected DEF cells were probed with rabbit-origin anti-HA monoclonal antibody (Abcam, Cambridge, UK), and the HA recombinant proteins were visualized using Cy3 goat anti-rabbit immunoglobulin (IgG; Jackson, West Grove, PA, USA). All probed cells were observed under a Leica SPE confocal microscope (Wetzlar, Germany).