sheep polyclonal igg  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: Inhibition of IL-34 Unveils Tissue-Selectivity and Is Sufficient to Reduce Microglial Proliferation in a Model of Chronic Neurodegeneration

doi: 10.3389/fimmu.2020.579000

Figure Lengend Snippet: Activation of the CSF1R signaling pathway by IL-34 and CSF-1 and inhibition of IL-34-mediated cell growth using IL-34 neutralizing antibodies. (A) N13 microglia cell line was stimulated with IL-34 (50 or 100 ng/mL), CSF-1 (50 or 100 ng/mL) or LPS (1 μg/mL) for 5 or 10 min. Cell lysates were subjected to Western blotting which showed increased phosphorylation of CSF1R and downstream ERK1/2 and AKT after IL-34 and CSF-1 stimulation. (B) IL-34 neutralizing antibodies used in this study (mouse monoclonal IgG2A [v1.1, ), rat monoclonal IgG2A (MAB5195, R&D Systems) and sheep polyclonal IgG (AF5195, R&D Systems)] prevented IL-34-dependent cell growth of M-NFS-60 in a similar nanomolar range. n = 3, data shown represent mean ± SEM, two-way ANOVA followed by Tukey's multiple comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001, comparisons are stimulations vs. unstimulated control (C) for each time point.

Article Snippet: Mouse myelogenous leukemia (M-NFS-60) cells were CSF-1 (R&D systems, 216-MC/CF) starved for 24 h. In white clear bottom 96-well-plates 10 μL IL-34 antibody (mouse monoclonal IgG2A (v1.1 manufactured by Genscript, ( , )), rat monoclonal IgG2A (MAB5195, R&D Systems) and sheep polyclonal IgG (AF5195, R&D Systems) and 10 μL IL-34 stimulus (R&D systems, 5195-ML-CF) were incubated at 37°C for 30 min before 80 μL M-NFS-60 cells (10 3 cells/well) were added.

Techniques: Activation Assay, Inhibition, Western Blot, Phospho-proteomics, Comparison, Control

Journal: Frontiers in Immunology

Article Title: Inhibition of IL-34 Unveils Tissue-Selectivity and Is Sufficient to Reduce Microglial Proliferation in a Model of Chronic Neurodegeneration

doi: 10.3389/fimmu.2020.579000

Figure Lengend Snippet: Intracerebral IL-34 antibody administration resulted in reduced microglia proliferation in ME7 prion mice. (A) Mice infected with prion disease received a single intracerebral injection of mouse- or human-specific anti-IL-34 (sheep polyclonal IgG) and brains were analyzed 1 week later. (B) Histological analysis of BrdU/Iba1-positive microglial cells in the cortex showed a reduction after treatment with a mouse-, but not with a human-specific antibody. Scale bar 100 μm. Naïve n = 6, ME7 n = 12, ME7 + isotype n = 10, ME7 + anti-mIL-34 n = 8, ME7 + anti-hIL-34 n = 7, data shown represent mean ± SEM, two-way ANOVA followed by Tukey's multiple comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Mouse myelogenous leukemia (M-NFS-60) cells were CSF-1 (R&D systems, 216-MC/CF) starved for 24 h. In white clear bottom 96-well-plates 10 μL IL-34 antibody (mouse monoclonal IgG2A (v1.1 manufactured by Genscript, ( , )), rat monoclonal IgG2A (MAB5195, R&D Systems) and sheep polyclonal IgG (AF5195, R&D Systems) and 10 μL IL-34 stimulus (R&D systems, 5195-ML-CF) were incubated at 37°C for 30 min before 80 μL M-NFS-60 cells (10 3 cells/well) were added.

Techniques: Infection, Injection, Comparison

Journal: Frontiers in Immunology

Article Title: Inhibition of IL-34 Unveils Tissue-Selectivity and Is Sufficient to Reduce Microglial Proliferation in a Model of Chronic Neurodegeneration

doi: 10.3389/fimmu.2020.579000

Figure Lengend Snippet: Activation of the CSF1R signaling pathway by IL-34 and CSF-1 and inhibition of IL-34-mediated cell growth using IL-34 neutralizing antibodies. (A) N13 microglia cell line was stimulated with IL-34 (50 or 100 ng/mL), CSF-1 (50 or 100 ng/mL) or LPS (1 μg/mL) for 5 or 10 min. Cell lysates were subjected to Western blotting which showed increased phosphorylation of CSF1R and downstream ERK1/2 and AKT after IL-34 and CSF-1 stimulation. (B) IL-34 neutralizing antibodies used in this study (mouse monoclonal IgG2A [v1.1, ), rat monoclonal IgG2A (MAB5195, R&D Systems) and sheep polyclonal IgG (AF5195, R&D Systems)] prevented IL-34-dependent cell growth of M-NFS-60 in a similar nanomolar range. n = 3, data shown represent mean ± SEM, two-way ANOVA followed by Tukey's multiple comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001, comparisons are stimulations vs. unstimulated control (C) for each time point.

Article Snippet: For acute treatment in ME7 prion mice, 1 μg of mouse or human-specific IL-34 neutralizing antibody (sheep polyclonal IgG, AF5195, or AF5265, R&D Systems) was stereotaxically and bilaterally injected into the dorsal hippocampus, coordinates from bregma: anteroposterior, ?2.0 mm; lateral, ±1.7 mm; depth, ?1.6 mm, 12 weeks after induction of prion disease.

Techniques: Activation Assay, Inhibition, Western Blot, Phospho-proteomics, Comparison, Control

Journal: Frontiers in Aging Neuroscience

Article Title: Studies on Colony Stimulating Factor Receptor-1 and Ligands Colony Stimulating Factor-1 and Interleukin-34 in Alzheimer's Disease Brains and Human Microglia

doi: 10.3389/fnagi.2017.00244

Figure Lengend Snippet: PCR primer sequences.

Article Snippet: Three different antibodies were used for detection of IL-34; a IL-34 sheep polyclonal (R&D Systems, Cat. No. AF5265), a rabbit polyclonal (Abcam, Cambridge, MA, Cat. No. ab75723), and a mouse monoclonal antibody (Abcam, Cat. No. ab101443).

Techniques: Amplification