IL-12/IL-35 p35 Antibody (27537)

Detection of IL‑12/IL‑35 p35 in PBMC's + LPS (1ug/mL) 24 hrs vs untreated PBMC's by Flow Cytometry

PBMC's + LPS (1ug/mL;24 hrs) and monensin (3 hrs) (A) vs untreated PBMC's (B) were stained with Mouse Anti-Human/Mouse IL‑12/IL‑35 p35 Monoclonal Antibody (Catalog # mab1570) and Mouse Anti-Human CD14 PE‑conjugated Monoclonal Antibody (Catalog # FAB3832P) followed by Allophycocyanin-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # F0101B). To facilitate intracellular staining, cells were fixed with Flow Cytometry Fixation Buffer (Catalog # FC004) and permeabilized with Flow Cytometry Permeabilization/Wash Buffer I (Catalog # FC005). View our protocol for Staining Intracellular Molecules.

Detection of IL-12/IL-35 p35 by Immunohistochemistry

IL-35 is highly expressed in retinal rod and cone photoreceptors. (A) qPCR analysis of p35, ebi3 IL-10 mRNA transcripts in the retina and spleen of WT mice. (B) Western blot analysis of whole cell extracts prepared from sorted CD4+ T cells, CD19+ B cells, dendritic cells (CD11c) and monocytes (Ly6C). Antibodies used were specific to mouse p35 or beta -actin. (C) Immunohistochemistry (IHC) analyses of frozen eye sections were performed with or without primary mouse IL-12p35 antibodies and goat anti-mouse (fab’2) secondary antibody. Slides were then stained with a DAB kit and counter-stained with methyl blue. IL-12p35-expressing cells, dark brown; microglia, white arrows; photoreceptor layers, red arrows. (D,E) Detection of IL-12p35-expressing cells by immunohistochemistry and confocal microscopy. Paraffin-fixed whole eye sections were subjected to antigen-retrieval, blocked in mouse blocking solution, incubated without or with primary rabbit IL-12p35 antibody and then goat anti-rabbit conjugated with AF647 antibody. Far-red magenta color indicates IL-12p35 expression; red arrow, photoreceptor layers; white arrow, microglial cells. GC, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL outer plexiform layer, ONL, outer nuclear layer; IS, inner section; OS, outer segment; RPE/choroid, retinal pigmented epithelial and choroid. Data represent at least 3 independent experiments. *** p < 0.001. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/35897732), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of IL-12/IL-35 p35 by Immunohistochemistry

IL-35 is highly expressed in retinal rod and cone photoreceptors. (A) qPCR analysis of p35, ebi3 IL-10 mRNA transcripts in the retina and spleen of WT mice. (B) Western blot analysis of whole cell extracts prepared from sorted CD4+ T cells, CD19+ B cells, dendritic cells (CD11c) and monocytes (Ly6C). Antibodies used were specific to mouse p35 or beta -actin. (C) Immunohistochemistry (IHC) analyses of frozen eye sections were performed with or without primary mouse IL-12p35 antibodies and goat anti-mouse (fab’2) secondary antibody. Slides were then stained with a DAB kit and counter-stained with methyl blue. IL-12p35-expressing cells, dark brown; microglia, white arrows; photoreceptor layers, red arrows. (D,E) Detection of IL-12p35-expressing cells by immunohistochemistry and confocal microscopy. Paraffin-fixed whole eye sections were subjected to antigen-retrieval, blocked in mouse blocking solution, incubated without or with primary rabbit IL-12p35 antibody and then goat anti-rabbit conjugated with AF647 antibody. Far-red magenta color indicates IL-12p35 expression; red arrow, photoreceptor layers; white arrow, microglial cells. GC, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL outer plexiform layer, ONL, outer nuclear layer; IS, inner section; OS, outer segment; RPE/choroid, retinal pigmented epithelial and choroid. Data represent at least 3 independent experiments. *** p < 0.001. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/35897732), licensed under a CC-BY license. Not internally tested by R&D Systems.