Figure 3

mAb-PN9cx3 is a broad neutralizing antibody against strains of both PRRSV-1 and PRRSV-2. A The PRRSV N transcript levels detected by RT-qPCR. PAMs were infected with PRRSV-SD16 virus at an MOI of 0.1 or PRRSV-SD16 virus pretreated (1-h incubation time) with 1 μM mAb-PN9cx3. After inoculation with the mAb-virus mixture, the PAMs were incubated for 1 h, and the unbound PRRSV virions were removed by washing the cells with fresh medium. The replication of PRRSV at 24 h after infection was determined by RT-qPCR to assess the PRRSV N protein-encoding transcript levels. The experiment was repeated three times. The significant differences between the mAb-PN9cx3-treated-and-infected groups and the PRRSV-infected only group (0 μM) are marked by asterisks: * (p < 0.05) and ** (p < 0.01). B Viral yields in cell culture supernatants of PAMs infected with different PRRSV strains or infected with PRRSV preincubated with 1 μM mAb-PN9cx3. Supernatants were collected at 24 hpi and titrated in MARC-145 cells. All above-described experiments were repeated at least three times. The significant differences between infected PAMs of the 0 μM mAb-PN9cx3 group and 1 μM mAb-PN9cx3 group are marked by asterisks: * (p < 0.05). C Western blotting of PAMs infected with PRRSV-GD-HD and JXA1 virus at an MOI of 0.1 or PRRSVs pretreated (1-h incubation time) with 1 μM mAb-PN9cx3. After inoculation with a mAb-virus mixture, the PAMs were incubated for 1 h, and unbound PRRSV virions were removed by washing the cells with fresh medium. The PAMs were then incubated for 24 h at 37 °C before lysis for SDS-PAGE and Western blot analysis of the PRRSV N protein levels. D Western blotting of PAMs infected with VR2332 and NADC30-like HNhx virus at an MOI of 0.1 or with PRRSVs pretreated (1-h incubation time) with 1 μM mAb-PN9cx3. After inoculation with a mAb-virus mixture, the PAMs were incubated for 1 h, and unbound PRRSV virions were removed by washing the cells with fresh medium. The PAMs were then incubated for 24 h at 37 °C before lysis for SDS-PAGE and Western blotting to determine the PRRSV N protein levels.