Analysis of recombinant PCV proteins.

<p>COS-7L cells were transiently transfected with pcDNA3.1-based plasmids encoding genes for the expression of PCV secreted prM/E, NS1 and partial NS5. Each expressed protein contained a C-terminal V5 tag. Mock cells were transfected with the empty pcDNA3.1 construct. <b>A</b>) Cells were fixed 21 hr post-transfection and probed with mAbs 3D6, 9G4 or anti-V5 using IFA. <b>B</b>) Lysates of COS-7L cells transfected with plasmids encoding PCV NS1, PCV secreted prM/E, WNV NS1 or pcDNA3.1 (M = mock) were treated (+) or untreated (−) with PNGaseF to remove N-linked glycans. The proteins were probed with anti-V5 (left panel) or anti-NS1 mAb 4G4 (right panel) in Western blot. <b>C,D</b>) Western blot analysis of supernatant and lysate of transiently transfected cells with PCV NS1 construct (<b>C</b>) or WNV NS1 construct (<b>D</b>). Protein detection was with anti-V5 and mAb 4G4 respectively. Lane 1 – NS1 lysate; Lane 2 – mock lysate; Lane 3 – NS1 supernatant; Lane 4 – mock supernatant. *Partial NS5 expressed. The construct is missing the sequence for the first 85 amino acids of the predicted full length PCV NS5 protein.