mouse monoclonal antibody against ox 6  (Abcam)

Journal: Frontiers in Molecular Neuroscience

Article Title: Upregulation of Cysteine Protease Cathepsin X in the 6-Hydroxydopamine Model of Parkinson’s Disease

doi: 10.3389/fnmol.2018.00412

Figure Lengend Snippet: Phenotype of increased cathepsin X-immunopositive cells in the lesioned SNc after 6-OHDA injection. Representative images of double immunofluorescence staining of cathepsin X (red fluorescence) and cell-type markers (green fluorescence) for neurons (NeuN, A ), microglial cells (OX-6, B ) and astrocytes (GFAP, C ) in the ipsilateral SNc at 12 h after the injection of 6-OHDA. Nuclei were counterstained with DAPI (blue fluorescence). Expression of cathepsin X in the lesioned SNc is predominantly restricted to NeuN-positive neuronal cells [ (A) arrows]. A single cathepsin X-positive microglial cell in the SNc was notable ( B , arrows), while most microglial cells were negative for cathepsin X (arrowheads). No astrocytes were positive for cathepsin X ( C , arrowheads). Groups of four animals ( n = 4) were analyzed and four sections of the SNc region of each animal were analyzed. Scale bar = 100 μm.

Article Snippet: They were then incubated overnight at 4°C with mouse monoclonal antibody against TH (1:750, Abcam, Cambridge, United Kingdom), goat polyclonal primary antibody against cathepsin X (1:200, AF934, R&D Systems, MN, United States) or mouse monoclonal antibody against OX-6 (1:300, Abcam, Cambridge, United Kingdom), diluted in blocking solution and then with biotinylated anti-mouse or anti-goat secondary antibodies (1:750, Vector Laboratories, Burlingame, CA, United States) diluted in KPBS containing 1% BSA and 0.02% Triton X-100 for 1.5 h at room temperature.

Techniques: Injection, Double Immunofluorescence Staining, Fluorescence, Expressing

Journal: Frontiers in Molecular Neuroscience

Article Title: Upregulation of Cysteine Protease Cathepsin X in the 6-Hydroxydopamine Model of Parkinson’s Disease

doi: 10.3389/fnmol.2018.00412

Figure Lengend Snippet: Phenotype of increased cathepsin X-immunopositive cells in the ipsilateral SNc after long-term 6-OHDA injection. (A) Enzymatic activity analysis of cathepsin X in the SNc, dissected from the coronal sections from the 4 weeks’ time-point after 6-OHDA-induced lesion. Significantly increased cathepsin X activity in the ipsilateral SNc (Lesion) was observed, compared to that in the contralateral side (Control). Values are means ± SD of four experiments. ∗ P < 0.05 (B–D) Representative images of double immunofluorescence staining of cathepsin X (red fluorescence) and cell-type markers (green fluorescence) for of neurons (NeuN, B ), microglial cells (OX-6, C ) or astrocytes (GFAP, D ) in the ipsilateral SNc at 4 weeks after the 6-OHDA injection. Nuclei were counterstained with DAPI (blue fluorescence). Neuronal cells surrounding the site of injection were negative for cathepsin X expression. Increased cathepsin X expression in ipsilateral SNc was predominantly restricted to microglial cells ( C , white arrows) and some astrocytes ( D , white arrows). Groups of four animals ( n = 4) were analyzed and four sections of the SNc region of each animal were analyzed. Scale bars = 100 μm.

Article Snippet: They were then incubated overnight at 4°C with mouse monoclonal antibody against TH (1:750, Abcam, Cambridge, United Kingdom), goat polyclonal primary antibody against cathepsin X (1:200, AF934, R&D Systems, MN, United States) or mouse monoclonal antibody against OX-6 (1:300, Abcam, Cambridge, United Kingdom), diluted in blocking solution and then with biotinylated anti-mouse or anti-goat secondary antibodies (1:750, Vector Laboratories, Burlingame, CA, United States) diluted in KPBS containing 1% BSA and 0.02% Triton X-100 for 1.5 h at room temperature.

Techniques: Injection, Activity Assay, Double Immunofluorescence Staining, Fluorescence, Expressing

Journal: Journal of Neuroinflammation

Article Title: Calcium dysregulation via L-type voltage-dependent calcium channels and ryanodine receptors underlies memory deficits and synaptic dysfunction during chronic neuroinflammation

doi: 10.1186/s12974-015-0262-3

Figure Lengend Snippet: Chronic LPS infusion increased activation of microglia but not astrocytes, and treatment with either dantrolene or nimodipine reduced microglia activation. Activated microglia were quantified by counting MHC-II+ cells, and activated astrocytes were quantified by densitometry of GFAP. Hippocampal immunohistochemistry against MHC-II/OX-6 (A) and GFAP was quantified (B, C, D, F, G, H) across specific hippocampal subfields. Although the DG region in LPS-infused rats contains numerous immunoreactive objects, only those OX-6 immunoreactive objects larger than 65 mm 2 were included in analysis and are represented in the histograms. GFAP hippocampal gene expression (E) was also quantified. (B) There was no change in the number of MHC-II+ cells in the CA1 subfield of the hippocampus. (C, D) There were significantly more MHC-II positive cells in the CA3 (C) and DG (D) subfield of all LPS-treated groups compared to their aCSF controls. Dantrolene treatment significantly reduced the number of MHC-II+ cells present in the CA3 and DG subfields, and nimodipine treatment significantly reduced the number of MHC-II+ cells present in the CA3. (E) There was a trend toward a drug × group interaction for hippocampal GFAP gene expression, but no statistically significant changes were observed. (F, G, H) There were no significant changes in the amount of GFAP in any of the hippocampal subfields due to either LPS or drug treatment. Data expressed as mean ± SEM. *Indicates a significant difference from treatment-matched aCSF controls, † Indicates a significant difference from LPS + vehicle rats. Significance determined by P < 0.05. LPS lipopolysaccharide, aCSF artificial cerebrospinal fluid, DG dentate gyrus, GFAP glial fibrillary acid protein.

Article Snippet: The rabbit polyclonal antibody against Arc (final dilution 1:2,000; Synaptic Systems, Goettingen, Germany) was used to label behaviorally activated neurons; the mouse monoclonal antibody against OX-6 (final dilution 1:200; BD Pharmigen, San Diego, CA, USA) was used to label major histocompatibility complex II (MHC-II) on activated microglia; and the mouse monoclonal antibody against glial fibrillary acid protein (GFAP; final dilution 1:2,000; Millipore Chemicon, Billerica, MA, USA) was used to quantify astrocyte activation.

Techniques: Activation Assay, Immunohistochemistry, Expressing