Journal: Immunology

Article Title: Identification of two subpopulations of purified human blood B cells, CD27- CD23+ and CD27high CD80+, that strongly express cell surface Toll-like receptor 9 and secrete high levels of interleukin-6.

doi: 10.1111/j.1365-2567.2008.02844.x

Figure Lengend Snippet: Figure 1. Cell-surface Toll-like receptor 9 (TLR9) expression on B cells sorted from peripheral blood mononuclear cells (PBMCs). (a) Western blot analysis of TLR9 protein expressed in human PBMC lysate (lane 1) and positive control (lane 2). The estimated molecular weight of TLR9 is 120 000; b-actin was used as the load- ing control. The Western blot was performed three times with simi- lar results. (b) TLR9 was detected by antibody labelling and flow cytometry analysis after gating for CD19+. (c) Summary of flow cytometry analysis of TLR9 expression by CD19+ B cells. The mean percentage of CD19+ B cells that were positive and negative for TLR9 expression is shown (mean ± SD from 10 independent experi- ments). (d) Detection of cell surface and intracellular TLR9 expres- sion by flow cytometry. Flow cytometry representations of TLR9 expression on B-cell surface and intracellular locations (one repre- sentative experiment out of three). Cells stained with the isotype control antibody provided the background level of TLR9 expression.

Article Snippet: The blots were then incubated with a primary mAb to TLR9 (clone 26C593; Imgenex) for 90 min at room temperature.

Techniques: Expressing, Western Blot, Positive Control, Molecular Weight, Control, Cytometry, Flow Cytometry, Staining

Journal: Immunology

Article Title: Identification of two subpopulations of purified human blood B cells, CD27- CD23+ and CD27high CD80+, that strongly express cell surface Toll-like receptor 9 and secrete high levels of interleukin-6.

doi: 10.1111/j.1365-2567.2008.02844.x

Figure Lengend Snippet: Figure 3. Detection of interleukin-6 (IL-6) in the culture superna- tants of the indicated B-cell subsets stimulated with oligodeoxydi- nucleotide (ODN; CpG 2006, CpG 2006-G5) or control ODN (CpG 2006c) for 48 hr. (a) CD19+ CD27) CD23+ TLR9+ and CD19+

Article Snippet: The blots were then incubated with a primary mAb to TLR9 (clone 26C593; Imgenex) for 90 min at room temperature.

Techniques: Control

Journal: PLoS ONE

Article Title: Recruitment of Cbl-b to B Cell Antigen Receptor Couples Antigen Recognition to Toll-Like Receptor 9 Activation in Late Endosomes

doi: 10.1371/journal.pone.0089792

Figure Lengend Snippet: (A) Representative confocal microscopic images of splenocytes from mice with indicated genotypes. For experiments, cells were stimulated through the BCR (green) for 30 minutes then fixed and stained for TLR9 (blue) and Lamp-1 (red)(n = 3). (B) Quantitation of experiments shown in (A) for fraction of cells demonstrating significant co-localization of TLR9 with Lamp-1 (n = 3, 30 cells/exp) (Cbl-b −/− vs. Cblb C373A or WT, p<0.001). (C) Quantitation of fractions of cells in (A) demonstrating significant co-localization between BCR and TLR9 (n = 3, 30 cells/exp). (D) In vitro assay of T-bet induction in response to ODN 1826 or control ODN targeted through the BCR (n = 3, p<0.01).

Article Snippet: Antibodies used for visualization were Lamp-1 (1D4B) (BD Biosciences), TLR9 (26C593.2) (Imgenex), Cathepsin L (CPLH-3G10) and Cbl-b (H-121 and H-454) (Santa Cruz Biotechnology).

Techniques: Staining, Quantitation Assay, In Vitro, Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: TLR2 and TLR9 synergistically control herpes simplex virus infection in the brain.

doi: 10.4049/jimmunol.181.12.8604

Figure Lengend Snippet: FIGURE 1. Induction of cytokines and chemokines in DC subtypes after stimulation with TLR ligands or HSV-2 infection. A–I, BM-DCs were generated from C57BL/6, TLR2/, TLR9/, TLR2/9/, and MyD88/ mice as described and treated with Pam3Csk4 (200 ng/ml), ODN1826 (1 M), and infected with HSV-2 strain 333 (3 106 PFU/ml) or UV-inactivated strain MS (3 106 PFU/ml). Conventional DCs (J–M) and pDCs (N–Q) were isolated from spleen cells as described, cultured, and infected with HSV-2 strain MS (3 106 PFU/ml). Supernatants were harvested 24 h posttreatment, IFN-/ was measured by bioassay, and IL-6, IL-12p40, and RANTES were measured by Luminex assay. Data are shown as means of measurements from triplicate cultures SEM.

Article Snippet: C57BL/6J, TLR2 / , TLR9 / , and TLR2/9 / mice were bred at Taconic M&B, and MyD88 mice were bred at the animal facility at the University of Copenhagen.

Techniques: Infection, Generated, Isolation, Cell Culture, Bioassay, Luminex

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: TLR2 and TLR9 synergistically control herpes simplex virus infection in the brain.

doi: 10.4049/jimmunol.181.12.8604

Figure Lengend Snippet: FIGURE 2. Induction of cytokines and che- mokines in peritoneal macrophages after HSV-2 infection. Peritoneal cells were harvested from C57BL/6, TLR2/, TLR9/, and TLR2/9/

Article Snippet: C57BL/6J, TLR2 / , TLR9 / , and TLR2/9 / mice were bred at Taconic M&B, and MyD88 mice were bred at the animal facility at the University of Copenhagen.

Techniques: Infection

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 1 TLR9 in situ hybridization on mouse lungs. Lung sections stained using TLR9 DIG-labeled RNA show staining (arrows) in several cell types (A). Lung sections from mice stained without a specific probe (B) lacks staining in any tissue. High magnification shows staining in vascular endo- thelium (arrowhead) (C), septal (arrow) as well as alveolar macrophages (chevron) (D), and bronchial epithelium (double arrow) (E). Scale bar: 100 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: In Situ Hybridization, Staining, Labeling

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 2 TLR9 immunohistochemistry on mouse lungs. Lung sections from mice stained with only a secondary antibody (A) lack staining in alveolar septa, whereas those stained with vWF antibody (B) shows staining in endothelium (arrowhead) alone. Lung sections stained using TLR9 antibody show staining in several tissues (arrows) (C), including bronchial epithelium (double arrow) (D), vascular endothelium (arrowhead) (E), and septal (arrow) as well as alveolar macrophages (chevron) (F). Scale bar: 100 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: Immunohistochemistry, Staining

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 3 TLR9 staining in a mouse neutrophil. TLR9 staining (arrows) observed in mouse lung neutrophil, epithelium (arrowhead), and endothelium (chevron). N, nucleus; AS, alveolar space. Scale bar: 4 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: Staining

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 4 TLR9 staining in a mouse type-II cell. TLR9 staining (arrows) observed in a mouse lung type-II cell. N, nucleus; AS, alveolar space; LB, lamellar bodies. Scale bar: 4 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: Staining

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 6 Human lung TLR9 in situ RNA hybridization. Lung sections from normal (A) or COPD (B) patients stained using TLR9 DIG-labeled RNA probe show staining (arrows) in several cell types with a clear influx of TLR9 positive cells in the bronchus. Staining without a RNA probe shows lack of staining in any tissue (C). High magnification shows staining in bronchial epithelium (double arrow) (E), vascular endothelium (arrowhead) (F), and septal (arrow) as well as alveolar (chevron) macrophages (G). Scale bar: 100 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: In Situ, Hybridization, Staining, Labeling

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 5 TLR9 staining in a mouse alveolar macrophage cell. TLR9 staining (arrows) observed in a mouse alveolar macrophage. N, nucleus; AS, alveolar space. Scale bar: 4 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: Staining

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 7 Human lung TLR9 immunohistochemistry. Lung sections from human patients stained with only a secondary antibody (A) lack staining in alveolar septa, whereas those stained with vWF antibody (B) show staining in endothelium (arrowhead) alone. Lung sections stained using TLR9 antibody show staining (arrows) in several cell types in both normal (C) as well as COPD patients (D). High magnification shows staining in bron- chial epithelium (double arrow) (E), vascular endothelium (arrowhead) (F), and septal (G) as well as alveolar macrophages (chevron) (H). Scale bar: 100 lm. Field counts of control and COPD patients showed a greater density of TLR9 positive cells in those with COPD (P < 0.01).

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: Immunohistochemistry, Staining, Control

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 8 TLR9 staining in human alveolar macrophage. TLR9 staining (arrows) observed in a human alveolar macrophage. N, nucleus; AS, alveolar space. Scale bar: 4 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: Staining

Journal: Journal of anatomy

Article Title: Expression of Toll-like receptor 9 in mouse and human lungs.

doi: 10.1111/joa.12039

Figure Lengend Snippet: Fig. 9 TLR9 staining in a human type-II cell. TLR9 staining (arrows) observed in a mouse lung type-II cell. N, nucleus; AS, alveolar space. Scale bar: 4 lm.

Article Snippet: Immuno-electron microscopy was carried out as described previously (Schneberger et al., 2009) with TLR9 antibody (1 : 50) and 20 nm gold-conjugated anti-mouse secondary antibody (1 : 100; Fitzgerald Industries International, Concord, MA, USA) for 1 h. The controls included protocols without the primary antibody and staining of sections with vWF antibody.

Techniques: Staining