Antibody Production and Characterization for Viral Proteins

Polyclonal antibodies (pAb) against E protein and prM protein of ZIKV, E protein of JEV, GP64, and VP39 of baculovirus were used as previously described [16 (link),17 (link),18 (link)]. The pAbs were purified by a Protein A-Sepharose^TM column (Sigma-Aidrich, Darmstadt, Germany). Mouse anti-Flavivirus group antigen antibody (2A10G6) was kindly provided by Dr. Chengfeng Qin (Beijing Institute of Microbiology and Epidemiology, Beijing, China). Mouse antibodies against HA, Strep, and V5-tag were purchased from Sigma (St. Louis, MO, USA). Goat anti-mouse and anti-rabbit IgG-fluorescein isothiocyanate (FITC; Abcam, Cambridge, UK) were used as the secondary antibodies in the immunofluorescence assays (IFA). Thiol-conjugating reagent 4-acetamido-4′-maleimidylstilbene-2,2′-disulfonic acid (AMS) was purchased from Invitrogen (Carlsbad, CA, USA). Proteasome inhibitor MG1332 was kindly provided by Dr. Leike Zhang (Wuhan Institute of Virology, Chinese Academy of Sciences).