Anti mmcp 8 antibody clone tug8

Excised eye specimens were fixed with 4% (wt/vol) paraformaldehyde and then embedded in paraffin. The tissues were sectioned at 4-μm thickness, and deparaffinized sections were subjected to hematoxylin and eosin (H&E) staining or immunohistochemistry. Immunofluorescence for IL-33 was as described previously^{7 (link)}. In brief, sections were incubated with an affinity-purified rabbit anti-mouse IL-33 polyclonal antibody, and bound antibodies were visualized with a biotinylated goat anti-rabbit IgG antibody (Vector Laboratories) and a Streptavidin, Alexa Fluor^® 594 conjugate (Invitrogen, Carlsbad, CA). Immunofluorescence for mMCP-8 was as described previously^{9 (link)}. Sections were incubated with anti-mMCP-8 antibody (clone TUG8, BioLegend), and bound antibodies were visualized with a Cy3-conjugated donkey anti-rat IgG antibody (WAKO, Osaka, Japan). Following mounting with a ProLong^® Diamond Antifade with DAPI (Life Technologies, Gaithersburg, MD), fluorescence images were recorded using a confocal laser scanning microscope LSM780 (Carl Zeiss MicroImaging, Thornwood, NY).

Sections of S. mansoni-infected or uninfected paraffin-embedded mouse tissues [liver and intestine/ileum from random-bred CD1 mice (Charles River, UK)] were stained for basophils with the basophil-specific rat anti-mMCP-8 antibody (clone TUG8, Biolegend), either by immunofluorescence or immunohistochemically. For immunofluorescence, anti-mMCP-8 was diluted 1:500. Binding was detected by AF-633-labeled secondary rabbit anti-rat IgG (Invitrogen), dilution 1:1000. Nuclei were counterstained by DAPI (Invitrogen; dilution 1:1,000), and pictures were taken with an Olympus IX-81 inverse fluorescence microscope (Olympus). Sections of S. mansoni-infected paraffin-embedded mouse liver tissue from female C57BL/6-NTAC mice (Taconic, Denmark) were immunohistochemically stained the anti-mMCP-8 and the secondary biotinylated rabbit anti-rat antibody were diluted 1:100, and detection was performed using the Vectastain Kit (Vector Laboratories). For detection of IPSE/alpha-1 the monoclonal antibody anti-IPSE/alpha-1 (clone 74 1G2) (16 (link)), was biotinylated according to standard protocols and diluted 1:100. To prevent non-specific binding to the mouse tissue, the M.O.M. Peroxidase kit (Vector laboratories) was applied. Mayer‘s hematoxylin (Merck) was used for counter staining. Pictures were taken at the Olympus BX51 microscope.