Journal: Graefe's Archive for Clinical and Experimental Ophthalmology

Article Title: Neural stem/progenitor cells circulating in peripheral blood of patients with neovascular form of AMD: a novel view on pathophysiology

doi: 10.1007/s00417-011-1767-9

Figure Lengend Snippet: Identification of CXCR4 + Lin – CD45 - TCSCs derived from the peripheral blood of AMD patients. The individual images depict the expression of CXCR4 and β-III-tubulin ( a ), GFAP ( b ) and nestin ( c ) antigens in isolated CXCR4 + Lin – CD45 - cells. The nuclei were visualized via DAPI staining. A pseudocolour was assigned to each stain as follows: anti-CXCR4 – red , anti-nestin, β-III-tubulin and GFAP – green , nuclei – blue . The control images (the upper boxed insets ) confirm no unspecific binding of antibodies. The scale bar shows 5 μm. Representative data are shown

Article Snippet: The sorted cells were permeabilized (0.1% Triton X-100, 10 min) and stained for β-III-tubulin (OriGene Technologies, Rockville, MD, USA), nestin (Novus Biologicals, Littleton, CO, USA) and glial fibrillary acidic protein (GFAP) (Santa Cruz Biotechnology, Santa Cruz, CA, USA) antigens with anti-human monoclonal antibodies for 1 h at room temperature followed by incubation with a secondary antibody conjugated to FITC (Sigma, St. Louis, MO, USA).

Techniques: Derivative Assay, Expressing, Isolation, Staining, Binding Assay

Journal: Graefe's Archive for Clinical and Experimental Ophthalmology

Article Title: Neural stem/progenitor cells circulating in peripheral blood of patients with neovascular form of AMD: a novel view on pathophysiology

doi: 10.1007/s00417-011-1767-9

Figure Lengend Snippet: Expression of nestin, β-III-tubulin, and GFAP at the mRNA level in peripheral blood nuclear cells. Transcript quantity is expressed in relative units against the β2-microglobulin control gene (2 ΔCt , where Ct represents the threshold cycle difference between the control and target genes)

Article Snippet: The sorted cells were permeabilized (0.1% Triton X-100, 10 min) and stained for β-III-tubulin (OriGene Technologies, Rockville, MD, USA), nestin (Novus Biologicals, Littleton, CO, USA) and glial fibrillary acidic protein (GFAP) (Santa Cruz Biotechnology, Santa Cruz, CA, USA) antigens with anti-human monoclonal antibodies for 1 h at room temperature followed by incubation with a secondary antibody conjugated to FITC (Sigma, St. Louis, MO, USA).

Techniques: Expressing

Journal: Graefe's Archive for Clinical and Experimental Ophthalmology

Article Title: Neural stem/progenitor cells circulating in peripheral blood of patients with neovascular form of AMD: a novel view on pathophysiology

doi: 10.1007/s00417-011-1767-9

Figure Lengend Snippet: Correlation between the expression of some neuroprogenitor and glial cell markers in the study groups

Article Snippet: The sorted cells were permeabilized (0.1% Triton X-100, 10 min) and stained for β-III-tubulin (OriGene Technologies, Rockville, MD, USA), nestin (Novus Biologicals, Littleton, CO, USA) and glial fibrillary acidic protein (GFAP) (Santa Cruz Biotechnology, Santa Cruz, CA, USA) antigens with anti-human monoclonal antibodies for 1 h at room temperature followed by incubation with a secondary antibody conjugated to FITC (Sigma, St. Louis, MO, USA).

Techniques: Expressing

Journal: Autophagy

Article Title: Autophagy inhibition promotes SNCA/alpha-synuclein release and transfer via extracellular vesicles with a hybrid autophagosome-exosome-like phenotype

doi: 10.1080/15548627.2017.1395992

Figure Lengend Snippet: ALP inhibition enhances SNCA release. (A, B) ICC showing increased expression and homogeneous SNCA distribution in SNCA-overexpressing H4 cells (SNCA H4 compared to CTR H4) and neurons (SNCA-tg compared to WT). TUBB3 or GFP were used to visualize cellular outline. Scale bar: 10 µm. SNCA expression was also examined by WB analysis of cell lysate from H4 cells (C) or neurons (D). SNCA is not detectable in CTR H4, therefore quantification was not applicable. SNCA-tg neurons overexpress the protein by 2.5 fold compared to WT (*p = 0.024, N = 3). ACTB or TUBB3 were used as loading controls. (E) Dot blot analysis of SNCA H4 conditioned medium (CM) shows that relative total extracellular SNCA levels increase by treatment with the ALP inhibitors Baf and CQ over the corresponding Veh, which is plotted as a dotted line. (For Baf: *p<0.05, with 2 nM N = 7, 10 nM N = 6, 20 and 200 nM N = 5, one sample t test; for 50 µM CQ: **p<0.008, N = 4, one sample t test.) (F) Total extracellular SNCA was assessed by ultra-sensitive ELISA, and values were normalized to total cell protein of the corresponding lysate. With 20 nM Baf or 50 µM CQ, an approximately 3-fold SNCA increase over Veh was measured (for Baf: **p = 0.002, N = 3; for CQ: *p = 0.013, N = 4). (G) The ratio of extracellular:intracellular SNCA was calculated from the ELISA measurements. Baf (20 nM) and CQ (50 µM) increased by approximately 4 fold the percent SNCA amount present extracellularly over intracellularly. (For Baf: ***p = 0.0003, N = 3; for CQ: ** p = 0.007, N = 4.)

Article Snippet: Nuclei were stained with 4’6’-diamidino-2-phenylindol (DAPI, 1:10,000; Sigma-Aldrich, D8417) for 5 min. SNCA (human) (15G7, 1:200; Alexis-Axxora, ALX-804-258) or SNCA (human and rodent) (Clone 42/α-Synuclein, 1:200; BD Transduction Laboratories, 610787), SQSTM1 (1:700; MBL, PM045), TUBB3 (1:1000; Promega, G7121), and MAP2 (Clone AP-20, 1:500; Sigma Aldrich, M1406) antibodies were used.

Techniques: Inhibition, Expressing, Dot Blot, Enzyme-linked Immunosorbent Assay