Quantifying Porcine Parvovirus Protein Levels

Western blotting was performed to compare loads of PRV proteins in plasma (0.4 μL) at 3 and 4 wpc. The proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), using 4–12% Bis-Tris Criterion XT PreCast gels (Bio-Rad, Hercules, CA, USA). The proteins were transferred to a polyvinylidene fluoride membrane (Trans-Blot Rurbo Midi PVDF, Bio-Rad, Hercules, CA, USA) and incubated with primary antibody overnight at 4 °C using the following sera diluted 1:500; Anti-σ1 #K275 [9 (link)], Anti-μ1C #K265 [9 (link)], Anti-λ1 #K273 [38 (link)] and Anti-σNS 01BO [39 (link)]. After incubation with secondary antibody, anti-rabbit IgG-HRP (Amersham, GE Healthcare, Buchinghamshire, UK) (1:50,000), the PRV proteins were detected by chemiluminescence (Clarity Western ECL Substrate, Bio-Rad, Hercules, CA, USA). MagicMark (XP Western Protein Standard, Invitrogen) was used as ladder. The western blot was performed on plasma samples at 3 and 4 wpc, detecting σ1 in all groups and expanded to include μ1C, λ1 and σNS in group NOR-2018/SF and CAN 16-005ND.