Journal: PLoS ONE

Article Title: Influenza A(H1N1)pdm09 Virus Suppresses RIG-I Initiated Innate Antiviral Responses in the Human Lung

doi: 10.1371/journal.pone.0049856

Figure Lengend Snippet: (A) The lung slices were processed for immunohistochemistry for detection of viral NP using rabbit polyclonal antibody (red). Panels a, b and c show mock (virus dilution buffer), OK/09 and PR8 infection, respectively. Panels d, e and f are corresponding bright-field images that demonstrate that lung architecture is preserved during the experiment. (B) Replication of influenza virus OK/09, OK/06 and PR8 in the human lung organ culture model. Lung slices exposed to virus at 6×10 6 PFU/ml were cultured for various times and total cellular mRNA was extracted. Quantitative RT-PCR was performed using Oligo dT as the primer for the first strand synthesis. Primers specific for NP were used to examine NP mRNA expression.

Article Snippet: Sections (3–5 µm) were mounted on glass slides and immuno-probed with a goat anti-human polyclonal antibody for IP-10 (R&D Systems), a rabbit anti-RIG-I polyclonal antibody (Abcam, Cambridge, MA), an anti-NP polyclonal antibody , an anti-CD 68 monoclonal antibody (Dakocytomation, Carpinteria CA) for macrophages or an anti-pan cytokeratin monoclonal antibody (Dakocytomation) for epithelial cells.

Techniques: Immunohistochemistry, Infection, Organ Culture, Cell Culture, Quantitative RT-PCR, Expressing

Journal: PLoS ONE

Article Title: Influenza A(H1N1)pdm09 Virus Suppresses RIG-I Initiated Innate Antiviral Responses in the Human Lung

doi: 10.1371/journal.pone.0049856

Figure Lengend Snippet: Lung slices were exposed to 6×10 6 PFU/ml of influenza virus PR8 or OK/09 or virus diluents for 24 h in the presence of brefeldin A (BFA) to enhance the detection of cytokines. Slices were then processed for immunohistochemistry for the detection of the chemokine IP-10 using goat polyclonal antibodies, viral nucleoprotein (NP) using rabbit polyclonal antibody, and macrophages using anti-CD68 monoclonal antibody. Nuclei were stained with SYTOX green. Top : OK/09. Bottom : PR8. A–D : fluorescent images that demonstrate nuclei ( A ; blue), NP ( B ; red), IP-10 ( C ; green), and macrophages ( D ; cyan). E : bright-field images that demonstrate that lung architecture is preserved during the experiment. F : overlays of the fluorescent images that demonstrate that influenza induces IP-10 in CD68 positive intraalveolar cells, likely alveolar macrophages (arrows). Bars = 100 µm.

Article Snippet: Sections (3–5 µm) were mounted on glass slides and immuno-probed with a goat anti-human polyclonal antibody for IP-10 (R&D Systems), a rabbit anti-RIG-I polyclonal antibody (Abcam, Cambridge, MA), an anti-NP polyclonal antibody , an anti-CD 68 monoclonal antibody (Dakocytomation, Carpinteria CA) for macrophages or an anti-pan cytokeratin monoclonal antibody (Dakocytomation) for epithelial cells.

Techniques: Immunohistochemistry, Staining

Journal: PLoS ONE

Article Title: Influenza A(H1N1)pdm09 Virus Suppresses RIG-I Initiated Innate Antiviral Responses in the Human Lung

doi: 10.1371/journal.pone.0049856

Figure Lengend Snippet: Lung slices were exposed to 6×10 6 PFU/ml of influenza virus PR8 or OK/09 or virus diluents for 24 h in the presence of BFA to enhance the detection of cytokines. Slices were then processed for immunohistochemistry for the detection of IP-10 using goat polyclonal antibodies, RIG-I using rabbit polyclonal antibody, and epithelial cells using anti-cytokeratin monoclonal antibody. Top : OK/09. Bottom : PR8. A–D : fluorescent images that demonstrate nuclei ( A ; blue), RIG-I ( B ; red), IP-10 ( C ; green), and epithelial cells ( D ; cyan). E : bright-field images. F : overlays of the fluorescent images that demonstrate that influenza induces RIG-I in epithelial cells (arrows). Bars = 100 µm.

Article Snippet: Sections (3–5 µm) were mounted on glass slides and immuno-probed with a goat anti-human polyclonal antibody for IP-10 (R&D Systems), a rabbit anti-RIG-I polyclonal antibody (Abcam, Cambridge, MA), an anti-NP polyclonal antibody , an anti-CD 68 monoclonal antibody (Dakocytomation, Carpinteria CA) for macrophages or an anti-pan cytokeratin monoclonal antibody (Dakocytomation) for epithelial cells.

Techniques: Immunohistochemistry