Figure 1 | Scientific Reports

Figure 1

From: RETRACTED ARTICLE: Neurotropic influenza A virus infection causes prion protein misfolding into infectious prions in neuroblastoma cells

Figure 1

IAV/WSN infection induces conversion of PrPC to PrPSc in N2aC24 cells. (a) Cell viability of N2aC24 cells after infection with different MOIs of IAV/WSN. Cell viability was determined by counting of live cells. (b) Western blotting of IAV/WSN-uninfected (Un) N2aC24 cells and -infected N2aC24 cells at various time points with 6D11 anti-PrP antibody, anti-HA antibody, anti-NP antibody, and anti-M2 antibody after treatment with ( +) or without (-) PK (20 μg/mg of proteins). PK-untreated 15 μg proteins from each cell lysate were used for detection of total PrP, HA NP, and M2 and PK-treated 300 μg proteins for PrPSc. (c) Left panels: Immunofluorescent staining of PrPSc (green) using 132 anti-PrP antibody in IAV/WSN-uninfected and -infected N2aC24 cells at 8 dpi and in 22L prion-infected N2aC24L1-3 cells. Right panel: The percentage (%) of 132 anti-PrP antibody-positive cells to DAPI-positive cells in at least 3 fields (145 × 109 μm/field) of IAV/WSN-uninfected (Un) and -infected N2aC24 cells at various time points and in 22L prion-infected N2aC24L1-3 (L1-3) cells. Blue, DAPI; bar, 10 μm. (d) Double immunofluorescent staining for PrPSc (green) and M2 (red) using 132 anti-PrP antibody and anti-HA antibody in IAV/WSN-uninfected and -infected N2aC24 cells at 8 dpi and in 22L prion-infected N2aC24L1-3 cells. Blue, DAPI; bar, 10 μm. (e) Left panel: Western blotting with 6D11 antibody of IAV/WSN-uninfected N2aC24 (C24) cells after 20 passages in IAV/WSN-free medium and N2aC24(R1) (C24R1) cells serially passaged after IAV/WSN infection using 15 μg proteins for total PrP (Left panel) and 30 μg proteins for PrPSc (Right panel). β-actin was used as an internal control. P, passages. (f) Western blotting of 15 μg proteins from N2aC24 and N2aC24(R1) cells with anti-HA antibody, anti-NP antibody, and anti-M2 antibody. IAV/WSN-infected N2aC24 cells were used as a positive control for the viral proteins. β-actin was used as an internal control. P, passages. Full length blots of Western blot images are shown in Supplementary Figs. 5c-e, 6, and 7.

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