Goat anti rabbit igg polymer dextran hrp

Detection of FCU1 and the viral proteins was performed by immunohistochemical labeling. For each treatment, 3 slides were analyzed. 5 days after IT injection of CPXtk⁻/gfp::fcu1 at the dose of 1 × 10⁶ pfu, resected LoVo tumors were fixed with 4% formaldehyde in 0.1 M phosphate buffer. Tumors where then desiccated and embedded in paraffin wax. Sections (5 μM) were mounted on adhesive glass slides and used for histological analysis.
Detection of FCU1 protein was performed by immunostaining of the slides fixed with tumor with anti-FCU1 mouse monoclonal antibody 3H1, followed by Goat anti Mouse-immunoglobulin-G (IgG)-Polymer Dextran HRP (DAKO, K4001) (red staining). CPXV-infected cells were detected upon incubation of the slide with rabbit IgG anti vaccinia-virus (B 65101R, Biodesign, dilution 1/1,000), followed by goat anti-rabbit IgG-Polymer Dextran HRP (DAKO, K4003, dilution 1/1,000) (green staining). To block non-specific antibody binding, slides were incubated 30 min with Linblock solution between the two staining steps. Coverslips were counterstained with DAPI (B-2883 SIGMA) (blue staining) and mounted on glass slides. Negative control tumors also underwent the same immunostaining treatment for comparison purposes. Slides were analyzed using Nikon microscopy.