Ifn α and ifn β

The stable HepG2-ISRE-Luc cells were seeded in 96-well plates that were either untransfected or transfected with corresponding constructs. After 24 h, the cells were treated for 2 h with the following antibodies: 20 μg/mL of a blocking α-IL10R2 antibody (R&D Systems), 40 μg/mL of a goat isotype IgG control (Abcam), a range (20, 10, 5, 2.5, or 1.25 μg/mL) of a custom rabbit monoclonal α-IFN-λ4 antibody (Prokunina-Olsson and others 2013 (link)), or 40 μg/mL of a rabbit isotype IgG control (Cell Signaling). Recombinant human interferons—10 ng/mL of custom IFN-λ3 (Prokunina-Olsson and others 2013 (link)), IFN-α and IFN-β (PBL Assay Science), or IFN-γ (R&D Systems) were added to the untransfected cells pretreated with the antibodies. Negative controls included nontreated cells, phosphate-buffered saline (PBS) in media, and mock transfection with control-Halo constructs. The cells were assayed for luciferase expression of the ISRE-Luc reporter 48 h post-transfection. All transfections and treatments were done in 8 biological replicates. For other experiments, inducible stable HepG2 cells were seeded into 96-well plates and protein expression was induced for 12 h. Antibodies (20 μg/mL of rabbit monoclonal α-IFN-λ4 or 40 μg/mL of rabbit IgG control) were added to shared culture media for 48 h.