Identification of a mAb that inhibits the biological activity of ECTV T1-IFNbp.

<p><b>A</b>) ELISA plates were covered with recombinant T1-IFNbp and the presence of the protein was detected with the indicated mAbs or isotype control (IC). Data are representatives of 2 independent experiments with similar results. <b>B</b>) L929 cells (10⁵) were incubated with 10 ng recombinant T1-IFNbp at 37°C for 1 h, thoroughly washed, and the binding of mAb 10G7 (blue line) or 10F3 (green) to the cell surface was analyzed by flow cytometry. <b>C</b>) L929 cells were infected with ECTV-GFP (upper panels) or ECTV-Δ166-GFP (lower panels) at MOI of 0.05 for 16 h. GFP expression and the binding of IC, 10G7 or 10F3 mAbs were determined by flow cytometry as indicated. <b>D</b>) 10 ng of recombinant ECTV T1-IFNbp was incubated with 10 µl rabbit anti-IFNbp or 10 ng of indicated mAbs for 30 min at 37°C, incubated with 1U mIFN-α for another 30 min at 37°C. The cocktail was then added to L929 cells in 96-well plates. The cells were incubated with this cocktail for 24 h, infected with VSV-eGFP at MOI of 0.1 for 16 h and observed for GFP fluorescence under the microscope.