recombinant human ocln  (Novus Biologicals)

Journal: Journal of Virology

Article Title: Splicing Diversity of the Human OCLN Gene and Its Biological Significance for Hepatitis C Virus Entry ▿ †

doi: 10.1128/JVI.00196-10

Figure Lengend Snippet: Splice forms of human OCLN have different abilities to mediate HCV entry. Human 786-O cells were mock transduced or transduced with human pTRIP-TagRFP-OCLN splice variants as indicated and then challenged in parallel with HCV and control VSV-G pseudoparticles (HCVpp and VSVGpp, respectively). HCVpp infectivity is reported as the titer of HCVpp divided by the titer of VSVGpp, after subtraction of the signals from infection with nonenveloped pseudoparticles (Env-pp). HCVpp infectivity is normalized to the signal in Huh-7.5 cells. Means and standard deviations from at least triplicate experiments are shown. wt, wild type.

Article Snippet: After being blocked with 5% milk for 1 h at room temperature, the membranes were incubated with appropriate primary antibodies (anti-OCLN H00004950-B01 [MaxPab; Abnova, Taiwan] raised against the full-length human OCLN protein [522 amino acids], mouse anti-α-tubulin [ab-7291; Abcam], and rabbit anti-HaloTag [Promega]), all at 1:1,000 dilutions, and then incubated with appropriate secondary IgG-horseradish peroxidase (HRP) antibodies (sc-2305 donkey anti-rabbit and sc-2302 goat anti-mouse; Santa Cruz), at 1:10,000 dilutions.

Techniques: Transduction, Control, Infection

Journal: Journal of Virology

Article Title: Splicing Diversity of the Human OCLN Gene and Its Biological Significance for Hepatitis C Virus Entry ▿ †

doi: 10.1128/JVI.00196-10

Figure Lengend Snippet: Expression of alternative splice forms of human OCLN does not significantly affect HCV entry. Human 786-O cells were mock transduced or (co)transduced with human Venus/YFP-WT-OCLN or TagRFP-OCLN splice variants as indicated and then challenged in parallel with HCV and control VSV-G pseudoparticles (HCVpp and VSVGpp, respectively). HCVpp infectivity is reported as the titer of HCVpp divided by the titer of VSVGpp, after subtraction of the signals from infection with nonenveloped pseudoparticles (Env-pp). HCVpp infectivity for the 786-O cell populations expressing the wild type or the splice variants at the indicated mean fluorescent intensities (MFIs) is normalized to the signal in Huh-7.5 cells. Means and standard deviations from at least triplicate experiments are shown. N/A, not applicable.

Article Snippet: After being blocked with 5% milk for 1 h at room temperature, the membranes were incubated with appropriate primary antibodies (anti-OCLN H00004950-B01 [MaxPab; Abnova, Taiwan] raised against the full-length human OCLN protein [522 amino acids], mouse anti-α-tubulin [ab-7291; Abcam], and rabbit anti-HaloTag [Promega]), all at 1:1,000 dilutions, and then incubated with appropriate secondary IgG-horseradish peroxidase (HRP) antibodies (sc-2305 donkey anti-rabbit and sc-2302 goat anti-mouse; Santa Cruz), at 1:10,000 dilutions.

Techniques: Expressing, Transduction, Control, Infection

Journal: Journal of Virology

Article Title: Splicing Diversity of the Human OCLN Gene and Its Biological Significance for Hepatitis C Virus Entry ▿ †

doi: 10.1128/JVI.00196-10

Figure Lengend Snippet: Genomic organization of the human 15q13.2 region. (A) The map shows the full-length OCLN gene with exons 1 to 9 and an inverted partly duplicated OCLN pseudogene (LOC647859) that includes exons 5 to 9. The HCV-binding MARVEL domain encoded by exons 3 and 4 of OCLN is underlined. (B) Structure of OCLN protein isoforms. Protein domains, exons, and translation start and stop sites are indicated. AA, amino acid.

Article Snippet: After being blocked with 5% milk for 1 h at room temperature, the membranes were incubated with appropriate primary antibodies (anti-OCLN H00004950-B01 [MaxPab; Abnova, Taiwan] raised against the full-length human OCLN protein [522 amino acids], mouse anti-α-tubulin [ab-7291; Abcam], and rabbit anti-HaloTag [Promega]), all at 1:1,000 dilutions, and then incubated with appropriate secondary IgG-horseradish peroxidase (HRP) antibodies (sc-2305 donkey anti-rabbit and sc-2302 goat anti-mouse; Santa Cruz), at 1:10,000 dilutions.

Techniques: Binding Assay

Journal: Journal of Virology

Article Title: Splicing Diversity of the Human OCLN Gene and Its Biological Significance for Hepatitis C Virus Entry ▿ †

doi: 10.1128/JVI.00196-10

Figure Lengend Snippet: OCLN protein isoforms

Article Snippet: After being blocked with 5% milk for 1 h at room temperature, the membranes were incubated with appropriate primary antibodies (anti-OCLN H00004950-B01 [MaxPab; Abnova, Taiwan] raised against the full-length human OCLN protein [522 amino acids], mouse anti-α-tubulin [ab-7291; Abcam], and rabbit anti-HaloTag [Promega]), all at 1:1,000 dilutions, and then incubated with appropriate secondary IgG-horseradish peroxidase (HRP) antibodies (sc-2305 donkey anti-rabbit and sc-2302 goat anti-mouse; Santa Cruz), at 1:10,000 dilutions.

Techniques:

Journal: Journal of Virology

Article Title: Splicing Diversity of the Human OCLN Gene and Its Biological Significance for Hepatitis C Virus Entry ▿ †

doi: 10.1128/JVI.00196-10

Figure Lengend Snippet: Protein expression of OCLN in human liver. Western blot showing expression pattern of OCLN protein isoforms in normal human liver samples. The observed OCLN protein isoforms are matched with their expected protein sizes (52.7 to 59.1, 31.6, and 23.3 kDa).

Article Snippet: After being blocked with 5% milk for 1 h at room temperature, the membranes were incubated with appropriate primary antibodies (anti-OCLN H00004950-B01 [MaxPab; Abnova, Taiwan] raised against the full-length human OCLN protein [522 amino acids], mouse anti-α-tubulin [ab-7291; Abcam], and rabbit anti-HaloTag [Promega]), all at 1:1,000 dilutions, and then incubated with appropriate secondary IgG-horseradish peroxidase (HRP) antibodies (sc-2305 donkey anti-rabbit and sc-2302 goat anti-mouse; Santa Cruz), at 1:10,000 dilutions.

Techniques: Expressing, Western Blot

Journal: Journal of Virology

Article Title: Splicing Diversity of the Human OCLN Gene and Its Biological Significance for Hepatitis C Virus Entry ▿ †

doi: 10.1128/JVI.00196-10

Figure Lengend Snippet: Analysis of subcellular localization of recombinant OCLN protein isoforms. (A) Confocal imaging of OCLN-HaloTag constructs transiently transfected into HeLa cells. Untransfected cells in the same field serve as negative controls. Antibody against α-tubulin was used to stain the cytoskeleton (green), while DAPI was used to stain the cell nuclei (blue). Images are obtained at 63× magnification with immersion oil. (B) Schematic representation of OCLN protein isoforms. EC1 and EC2 mark extracellular parts of the OCLN protein.

Article Snippet: After being blocked with 5% milk for 1 h at room temperature, the membranes were incubated with appropriate primary antibodies (anti-OCLN H00004950-B01 [MaxPab; Abnova, Taiwan] raised against the full-length human OCLN protein [522 amino acids], mouse anti-α-tubulin [ab-7291; Abcam], and rabbit anti-HaloTag [Promega]), all at 1:1,000 dilutions, and then incubated with appropriate secondary IgG-horseradish peroxidase (HRP) antibodies (sc-2305 donkey anti-rabbit and sc-2302 goat anti-mouse; Santa Cruz), at 1:10,000 dilutions.

Techniques: Recombinant, Imaging, Construct, Transfection, Staining