FIG 9.

Occurrence of ORF3 protein in PEDV virions. Vero cells were mock infected or infected with PEDVs at an MOI of 2 in T225 flasks. After 2 days of incubation, the cells were harvested and viral particles in the culture medium were purified by sucrose gradient centrifugation. Both the cellular samples and purified virions were analyzed for their protein compositions in a separate SDS-10% polyacrylamide gel which was subsequently processed for immunoblot analysis with antibodies against ORF3 (rabbit anti-ORF3 polyclonal antibody, P71-3; 1:2,000) and N (mouse anti-PEDV N protein monoclonal antibody, 3F-12; 1:2,000) proteins; GAPDH served as a protein loading control. Lanes corresponding with the lysates of the cells (C), the corresponding culture media (M), and the purified virus particles are indicated. The rightmost lane, labeled p71, refers to the analysis of an ORF3-glutathione S-transferase (GST) positive-control protein, derived by prokaryotic expression of pGEX-4T-1-p71, which specifies a fusion protein consisting of ORF3 aa 1 to 71 linked to a GST tag (∼33.5 kDa).