il 10 fitc  (Thermo Fisher)


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    Name:
    IL 10 Monoclonal Antibody BT 10
    Description:
    IL 10 Monoclonal Antibody for ELISA
    Catalog Number:
    BMS131-2
    Price:
    None
    Category:
    Antibodies Secondary Detection Reagents
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    Structured Review

    Thermo Fisher il 10 fitc
    Rebamipide reciprocally regulates Th17/Treg cell imbalance. Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. a The tissues were stained with specific Abs against CD4 ( green ), CD25 ( white ), IL-17 ( red ), FOXP3 ( green ), p -STAT3–Y705 ( red ), and p -STAT3–S727 ( red ). Positive cells are shown in the bar graphs . b Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. Splenocytes were stained with Abs against CD4–PerCP, IL-17–PE, and <t>IL-10–FITC</t> to determine the presence of Th17 and IL-10-expressing Treg cells. Data are expressed as the mean ± SEM of 3 independent experiments (* P
    IL 10 Monoclonal Antibody for ELISA
    https://www.bioz.com/result/il 10 fitc/product/Thermo Fisher
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    il 10 fitc - by Bioz Stars, 2021-06
    93/100 stars

    Images

    1) Product Images from "Rebamipide prevents peripheral arthritis and intestinal inflammation by reciprocally regulating Th17/Treg cell imbalance in mice with curdlan-induced spondyloarthritis"

    Article Title: Rebamipide prevents peripheral arthritis and intestinal inflammation by reciprocally regulating Th17/Treg cell imbalance in mice with curdlan-induced spondyloarthritis

    Journal: Journal of Translational Medicine

    doi: 10.1186/s12967-016-0942-5

    Rebamipide reciprocally regulates Th17/Treg cell imbalance. Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. a The tissues were stained with specific Abs against CD4 ( green ), CD25 ( white ), IL-17 ( red ), FOXP3 ( green ), p -STAT3–Y705 ( red ), and p -STAT3–S727 ( red ). Positive cells are shown in the bar graphs . b Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. Splenocytes were stained with Abs against CD4–PerCP, IL-17–PE, and IL-10–FITC to determine the presence of Th17 and IL-10-expressing Treg cells. Data are expressed as the mean ± SEM of 3 independent experiments (* P
    Figure Legend Snippet: Rebamipide reciprocally regulates Th17/Treg cell imbalance. Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. a The tissues were stained with specific Abs against CD4 ( green ), CD25 ( white ), IL-17 ( red ), FOXP3 ( green ), p -STAT3–Y705 ( red ), and p -STAT3–S727 ( red ). Positive cells are shown in the bar graphs . b Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. Splenocytes were stained with Abs against CD4–PerCP, IL-17–PE, and IL-10–FITC to determine the presence of Th17 and IL-10-expressing Treg cells. Data are expressed as the mean ± SEM of 3 independent experiments (* P

    Techniques Used: Isolation, Mouse Assay, Injection, Staining, Expressing

    Related Articles

    Recombinant:

    Article Title: Oral antigen exposure in newborn piglets circumvents induction of oral tolerance in response to intraperitoneal vaccination in later life
    Article Snippet: .. The reagents were as follows: Coating antibody: monoclonal anti-swine IL-10 (Invitrogen ASC0104), Detection antibody: monoclonal anti-swine IL-10 biotin (Invitrogen ASC9109), Standard: recombinant swine IL-10 (Invitrogen PSC0104) and Coating antibody: monoclonal anti-swine IFN-γ (Fisher Scientific Ltd, ENMP700), Detection antibody: monoclonal anti-swine IFN-γ (Fisher Scientific Ltd,ENPP700; biotinylated in-house), Standard: recombinant swine IFN-γ (Ceiba Geigy). .. The multiplex assay was carried out in a 96 well Grenier Bio-One Fluotrac 200 96 F black (VWR CanLab Mississauga, ON, #82050-754) which allows washing and retention of the Luminex beads.

    Staining:

    Article Title: IL-1β production by intermediate monocytes is associated with immunopathology in cutaneous leishmaniasis
    Article Snippet: For ex-vivo cell surface staining, incubated cells with monoclonal antibodies anti-CD14 (APC) and anti-CD16 (PE) (BD Pharmingen) or (FITC) (eBioscience), for 15 minutes, washed by centrifugation twice and fixed with 2% paraformaldehyde. .. For intracellular staining, cells were ressuspended in Perm/Wash (BD Biosciences) for 15 minutes and intracellular labeling was performed using monoclonal antibody anti-IL-1β (FITC) (eBioscience) and anti-NLRP3 (PE) (R & D Systems) for 30 minutes. .. Monocyte-derived macrophages were prepared following a method previously shown ( ) by our laboratory to yield 99% macrophages characterized by flow cytometry as CD14-positive, CD3-negative, CD19-negative.

    Article Title: Overexpression of soluble RAGE in mesenchymal stem cells enhances their immunoregulatory potential for cellular therapy in autoimmune arthritis
    Article Snippet: Confocal microscopy and immunostaining Tissues were obtained 42 days after CII immunization, snap-frozen in liquid nitrogen, and stored at −80 °C. .. Tissue cryosections (7 μm thick) were fixed in 4% (v/v) paraformaldehyde and stained using fluorescein isothiocyanate (FITC)-, phycoerythrin-, PerCP–Cy5.5-, or allophycocyanin-conjugated monoclonal antibodies to IL-10, CD4, CD25, IL-17, Foxp3, pSTAT-3 (Y705 and S727), pSTAT-5 (Y694) (all from eBioscience, San Diego, CA, USA), IDO (BD Biosciences, San Jose, CA, USA), and DAPI. .. After overnight incubation at 4 °C, the stained sections were visualized by confocal microscopy (LSM 510 Meta; Zeiss, Göttingen, Germany).

    Article Title: Rapamycin-treated human endothelial cells preferentially activate allogeneic regulatory T cells
    Article Snippet: For intracellular cytokine staining, T cells cocultured with allogeneic ECs for 6 days were stimulated for 6 hours with 40 ng/ml PMA and 2 μM ionomycin in the presence of Brefeldin A (eBioscience). .. Stimulated T cells were surface stained with anti–human CD4, CD25, CD127 fluorescent antibodies, then permeabilized using FoxP3 Staining Kit and stained with FoxP3-APC and IL-17–FITC (eBioscience), IL-10–FITC (eBioscience), IFN-γ–FITC (eBioscience), or TGF-β/LAP–FITC (BioLegend). .. To ensure antibodies were functional, positive staining was analyzed in the following controls: CD14+ monocytes stimulated with LPS (IL-10), CD4+ T cells stimulated by MHC II+ ECs (IFN-γ and IL-17), and CD4+ T cells stimulated with anti-CD3 and anti-CD28 in the presence of IL-2 (LAP).

    Article Title: Rebamipide prevents peripheral arthritis and intestinal inflammation by reciprocally regulating Th17/Treg cell imbalance in mice with curdlan-induced spondyloarthritis
    Article Snippet: Populations of T cells were examined by staining the tissues with a monoclonal Ab against CD4–peridin chlorophyll protein (eBioscience). .. Cells were permeabilized and fixed with CytoFix (BD Biosciences, San Jose, CA, USA), as instructed by the manufacturer, and were stained with Abs against IL-17–PE and IL-10–FITC (eBioscience). ..

    Labeling:

    Article Title: IL-1β production by intermediate monocytes is associated with immunopathology in cutaneous leishmaniasis
    Article Snippet: For ex-vivo cell surface staining, incubated cells with monoclonal antibodies anti-CD14 (APC) and anti-CD16 (PE) (BD Pharmingen) or (FITC) (eBioscience), for 15 minutes, washed by centrifugation twice and fixed with 2% paraformaldehyde. .. For intracellular staining, cells were ressuspended in Perm/Wash (BD Biosciences) for 15 minutes and intracellular labeling was performed using monoclonal antibody anti-IL-1β (FITC) (eBioscience) and anti-NLRP3 (PE) (R & D Systems) for 30 minutes. .. Monocyte-derived macrophages were prepared following a method previously shown ( ) by our laboratory to yield 99% macrophages characterized by flow cytometry as CD14-positive, CD3-negative, CD19-negative.

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    Thermo Fisher il 10 fitc
    Rebamipide reciprocally regulates Th17/Treg cell imbalance. Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. a The tissues were stained with specific Abs against CD4 ( green ), CD25 ( white ), IL-17 ( red ), FOXP3 ( green ), p -STAT3–Y705 ( red ), and p -STAT3–S727 ( red ). Positive cells are shown in the bar graphs . b Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. Splenocytes were stained with Abs against CD4–PerCP, IL-17–PE, and <t>IL-10–FITC</t> to determine the presence of Th17 and IL-10-expressing Treg cells. Data are expressed as the mean ± SEM of 3 independent experiments (* P
    Il 10 Fitc, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/il 10 fitc/product/Thermo Fisher
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    il 10 fitc - by Bioz Stars, 2021-06
    93/100 stars
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    94
    Thermo Fisher fitc anti il 10
    Flow cytometry analysis of the numbers of circulating <t>IL-10</t> + B cells. PBMCs from individual subjects were plated in 24-well plates (10 6 cells/well) and stimulated in duplicate with 50 ng/mL of PMA, 1.0 μg/mL of ionomycin, and 50 ng/mL of LPS in complete RPMI-1640 medium for 2 h at 37°C in 5% CO 2 followed exposing to Brefeldin A for another 4 h. The cells were then harvested, washed with ice-cold PBS, and stained with APC-anti-CD19, PerCP-anti-CD5, and PE-anti-CD1d. The immunostained cells were fixed and permeabilized with 0.5% saponin. After being washed, the cells were stained intracellularly with <t>FITC-anti-IL-10</t> and analyzed by flow cytometry. The cells were first gated on living lymphocytes and then on CD19 + B cells for further analysis of total CD19 + B cells, CD5 + CD19 + , CD5 + CD19 + CD1d high B cells, CD5 + CD19 + CD1 dhigh IL-10 + Bregs. The isotype-matched antibodies served as controls. The levels of serum IL-10 in individual subjects were analyzed by ELISA. Data are representative FACS charts and expressed as the mean values of individual subjects and the difference between groups was analyzed by the Kruskal–Wallis test. (A) Flow cytometry analysis; (B) quantitative analysis of the numbers of total CD19 + B cells, CD5 + CD19 + , CD5 + CD19 + CD1d high B cells, CD5 + CD19 + CD1d high IL-10 + Bregs and the levels of serum IL-10. The horizontal lines indicate the median values for different groups.
    Fitc Anti Il 10, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fitc anti il 10/product/Thermo Fisher
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    fitc anti il 10 - by Bioz Stars, 2021-06
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    99
    Thermo Fisher pe labeled anti il 22
    Supernatants of 1,25D diff -DCs promote differentiation of <t>IL-22-secreting</t> T cells. Monocytes were differentiated into DCs with rGM-CSF and rIL-4 in media with 10% FCS in the presence (1,25D diff -DCs) or absence (serum-DCs) of additional 1,25D (10 −8 M). 1,25D diff -DCs and serum-DCs were stimulated with TLR2/1L (1 μg/ml) in fresh media and supernatants were collected after 18–24 hours. Subsequently, TLR2/1-induced 1,25D diff -DC and serum-DC supernatants were added to naïve CD4 + T cells activated with CD3/CD28-coated beads. As a control, naïve CD4 + T cells were incubated with CD3/CD28-coated beads without addition of DC supernatants (beads only). After five days, rIL2 was added to all cultures. On day 12, T cells were re-stimulated with PMA/Ionomycin in fresh media and cytokine secretion evaluated after 18–24 hours. Levels of T cell-derived IL-22, IL-17a and IFN-γ were assessed by ELISA (mean of cytokine levels in ng/ml ± SEM, IL-22 n = 15, IL-17a/IFN-γ n = 13). *p
    Pe Labeled Anti Il 22, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pe labeled anti il 22/product/Thermo Fisher
    Average 99 stars, based on 1 article reviews
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    95
    Thermo Fisher anti mouse il 10 fitc
    The effect of ethyl pyruvate (EP) on Treg in vitro. CD4 + CD25 − T lymphocytes were stimulated for 24 h with a conventional Treg differentiation cocktail (control) and then additionally treated with 125 μM EP. ( A ) Treg (CD4 + CD25 high FoxP3 + ) proportion, at indicated time points after the addition of EP. Representative dot plots for the 72nd h are shown on the right-hand side. ( B ) Carboxyfluorescein succinimidyl ester (CFSE)-based proliferation of Treg (CD4 + CD25 high ) was evaluated by flow cytometry (divided, Div. and undivided, Und. cells, with representative histogram plots shown on the right-hand side) 48 h after EP treatment. Unstimulated cells were stained with CFSE and served as the control for setting the threshold for undivided cells. ( C ) The proportion of Treg expressing perforin, granzyme or CD39, 72 h after EP treatment. Representative dot plots for perforin + Treg are shown on the right-hand side. ( D ) mRNA expression of Tgf-β and <t>Il-10</t> in cultures treated with or without EP. * p
    Anti Mouse Il 10 Fitc, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti mouse il 10 fitc/product/Thermo Fisher
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    Image Search Results


    Rebamipide reciprocally regulates Th17/Treg cell imbalance. Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. a The tissues were stained with specific Abs against CD4 ( green ), CD25 ( white ), IL-17 ( red ), FOXP3 ( green ), p -STAT3–Y705 ( red ), and p -STAT3–S727 ( red ). Positive cells are shown in the bar graphs . b Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. Splenocytes were stained with Abs against CD4–PerCP, IL-17–PE, and IL-10–FITC to determine the presence of Th17 and IL-10-expressing Treg cells. Data are expressed as the mean ± SEM of 3 independent experiments (* P

    Journal: Journal of Translational Medicine

    Article Title: Rebamipide prevents peripheral arthritis and intestinal inflammation by reciprocally regulating Th17/Treg cell imbalance in mice with curdlan-induced spondyloarthritis

    doi: 10.1186/s12967-016-0942-5

    Figure Lengend Snippet: Rebamipide reciprocally regulates Th17/Treg cell imbalance. Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. a The tissues were stained with specific Abs against CD4 ( green ), CD25 ( white ), IL-17 ( red ), FOXP3 ( green ), p -STAT3–Y705 ( red ), and p -STAT3–S727 ( red ). Positive cells are shown in the bar graphs . b Spleen tissues were isolated from mice in each group at 14 weeks after curdlan injection. Splenocytes were stained with Abs against CD4–PerCP, IL-17–PE, and IL-10–FITC to determine the presence of Th17 and IL-10-expressing Treg cells. Data are expressed as the mean ± SEM of 3 independent experiments (* P

    Article Snippet: Cells were permeabilized and fixed with CytoFix (BD Biosciences, San Jose, CA, USA), as instructed by the manufacturer, and were stained with Abs against IL-17–PE and IL-10–FITC (eBioscience).

    Techniques: Isolation, Mouse Assay, Injection, Staining, Expressing

    Flow cytometry analysis of the numbers of circulating IL-10 + B cells. PBMCs from individual subjects were plated in 24-well plates (10 6 cells/well) and stimulated in duplicate with 50 ng/mL of PMA, 1.0 μg/mL of ionomycin, and 50 ng/mL of LPS in complete RPMI-1640 medium for 2 h at 37°C in 5% CO 2 followed exposing to Brefeldin A for another 4 h. The cells were then harvested, washed with ice-cold PBS, and stained with APC-anti-CD19, PerCP-anti-CD5, and PE-anti-CD1d. The immunostained cells were fixed and permeabilized with 0.5% saponin. After being washed, the cells were stained intracellularly with FITC-anti-IL-10 and analyzed by flow cytometry. The cells were first gated on living lymphocytes and then on CD19 + B cells for further analysis of total CD19 + B cells, CD5 + CD19 + , CD5 + CD19 + CD1d high B cells, CD5 + CD19 + CD1 dhigh IL-10 + Bregs. The isotype-matched antibodies served as controls. The levels of serum IL-10 in individual subjects were analyzed by ELISA. Data are representative FACS charts and expressed as the mean values of individual subjects and the difference between groups was analyzed by the Kruskal–Wallis test. (A) Flow cytometry analysis; (B) quantitative analysis of the numbers of total CD19 + B cells, CD5 + CD19 + , CD5 + CD19 + CD1d high B cells, CD5 + CD19 + CD1d high IL-10 + Bregs and the levels of serum IL-10. The horizontal lines indicate the median values for different groups.

    Journal: Journal of Translational Medicine

    Article Title: Increased numbers of CD5+CD19+CD1dhighIL-10+ Bregs, CD4+Foxp3+ Tregs, CD4+CXCR5+Foxp3+ follicular regulatory T (TFR) cells in CHB or CHC patients

    doi: 10.1186/s12967-014-0251-9

    Figure Lengend Snippet: Flow cytometry analysis of the numbers of circulating IL-10 + B cells. PBMCs from individual subjects were plated in 24-well plates (10 6 cells/well) and stimulated in duplicate with 50 ng/mL of PMA, 1.0 μg/mL of ionomycin, and 50 ng/mL of LPS in complete RPMI-1640 medium for 2 h at 37°C in 5% CO 2 followed exposing to Brefeldin A for another 4 h. The cells were then harvested, washed with ice-cold PBS, and stained with APC-anti-CD19, PerCP-anti-CD5, and PE-anti-CD1d. The immunostained cells were fixed and permeabilized with 0.5% saponin. After being washed, the cells were stained intracellularly with FITC-anti-IL-10 and analyzed by flow cytometry. The cells were first gated on living lymphocytes and then on CD19 + B cells for further analysis of total CD19 + B cells, CD5 + CD19 + , CD5 + CD19 + CD1d high B cells, CD5 + CD19 + CD1 dhigh IL-10 + Bregs. The isotype-matched antibodies served as controls. The levels of serum IL-10 in individual subjects were analyzed by ELISA. Data are representative FACS charts and expressed as the mean values of individual subjects and the difference between groups was analyzed by the Kruskal–Wallis test. (A) Flow cytometry analysis; (B) quantitative analysis of the numbers of total CD19 + B cells, CD5 + CD19 + , CD5 + CD19 + CD1d high B cells, CD5 + CD19 + CD1d high IL-10 + Bregs and the levels of serum IL-10. The horizontal lines indicate the median values for different groups.

    Article Snippet: After being washed, the cells were stained intracellularly with FITC-anti-IL-10 (eBiosciences) and analyzed by flow cytometry.

    Techniques: Flow Cytometry, Cytometry, Staining, Enzyme-linked Immunosorbent Assay, FACS

    Supernatants of 1,25D diff -DCs promote differentiation of IL-22-secreting T cells. Monocytes were differentiated into DCs with rGM-CSF and rIL-4 in media with 10% FCS in the presence (1,25D diff -DCs) or absence (serum-DCs) of additional 1,25D (10 −8 M). 1,25D diff -DCs and serum-DCs were stimulated with TLR2/1L (1 μg/ml) in fresh media and supernatants were collected after 18–24 hours. Subsequently, TLR2/1-induced 1,25D diff -DC and serum-DC supernatants were added to naïve CD4 + T cells activated with CD3/CD28-coated beads. As a control, naïve CD4 + T cells were incubated with CD3/CD28-coated beads without addition of DC supernatants (beads only). After five days, rIL2 was added to all cultures. On day 12, T cells were re-stimulated with PMA/Ionomycin in fresh media and cytokine secretion evaluated after 18–24 hours. Levels of T cell-derived IL-22, IL-17a and IFN-γ were assessed by ELISA (mean of cytokine levels in ng/ml ± SEM, IL-22 n = 15, IL-17a/IFN-γ n = 13). *p

    Journal: PLoS ONE

    Article Title: Vitamin D Regulates Cytokine Patterns Secreted by Dendritic Cells to Promote Differentiation of IL-22-Producing T Cells

    doi: 10.1371/journal.pone.0130395

    Figure Lengend Snippet: Supernatants of 1,25D diff -DCs promote differentiation of IL-22-secreting T cells. Monocytes were differentiated into DCs with rGM-CSF and rIL-4 in media with 10% FCS in the presence (1,25D diff -DCs) or absence (serum-DCs) of additional 1,25D (10 −8 M). 1,25D diff -DCs and serum-DCs were stimulated with TLR2/1L (1 μg/ml) in fresh media and supernatants were collected after 18–24 hours. Subsequently, TLR2/1-induced 1,25D diff -DC and serum-DC supernatants were added to naïve CD4 + T cells activated with CD3/CD28-coated beads. As a control, naïve CD4 + T cells were incubated with CD3/CD28-coated beads without addition of DC supernatants (beads only). After five days, rIL2 was added to all cultures. On day 12, T cells were re-stimulated with PMA/Ionomycin in fresh media and cytokine secretion evaluated after 18–24 hours. Levels of T cell-derived IL-22, IL-17a and IFN-γ were assessed by ELISA (mean of cytokine levels in ng/ml ± SEM, IL-22 n = 15, IL-17a/IFN-γ n = 13). *p

    Article Snippet: For intracellular cytokine staining the following monoclonal antibodies were used: PE-labeled anti-IL-22 (clone 22URTI, eBioscience), PerCP-Cy5.5-labeled anti-IFN-γ (clone B27, BD Biosciences), Alexa Fluor 647-labeled anti-IL-17a (clone N49-653, BD Biosciences), APC-labeled anti-IL-4 (clone 8D4-8, BD Biosciences).

    Techniques: Incubation, Derivative Assay, Enzyme-linked Immunosorbent Assay

    Supernatants of 1,25D diff -DCs promote differentiation of IL-22-expressing T cells. Supernatants of TLR2/1-induced 1,25D diff -DCs and serum-DC were added to naïve CD4 + T cells activated with CD3/CD28-coated beads (as described in Fig 4 ). After five days, rIL2 was added to all cultures. On day 12, T cells were re-stimulated with PMA/Ionomycin for five hours, the last 2.5 hours of culture in the presence of Brefeldin A, in fresh media and intracellular cytokine expression of IL-22, IFN-γ or IL-17a was measured. (A) Dot plots from one representative staining of one donor out of eleven. Upper panel of dot plots shows co-expression of IL-17a and IL-22, lower panel shows co-expression of IFN-γ and IL-22. Numbers above each dot plot indicate frequency of positive cells in each quadrant. (B) Frequency of total IL-22-, IL-17a- and IFN-γ-expressing CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 11). (C) Frequency of IL-22 + /IL-17a + and IL-22 + /IL-17a - or IL-22 + /IFN-γ + and IL-22 + /IFN-γ - CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 11). *p

    Journal: PLoS ONE

    Article Title: Vitamin D Regulates Cytokine Patterns Secreted by Dendritic Cells to Promote Differentiation of IL-22-Producing T Cells

    doi: 10.1371/journal.pone.0130395

    Figure Lengend Snippet: Supernatants of 1,25D diff -DCs promote differentiation of IL-22-expressing T cells. Supernatants of TLR2/1-induced 1,25D diff -DCs and serum-DC were added to naïve CD4 + T cells activated with CD3/CD28-coated beads (as described in Fig 4 ). After five days, rIL2 was added to all cultures. On day 12, T cells were re-stimulated with PMA/Ionomycin for five hours, the last 2.5 hours of culture in the presence of Brefeldin A, in fresh media and intracellular cytokine expression of IL-22, IFN-γ or IL-17a was measured. (A) Dot plots from one representative staining of one donor out of eleven. Upper panel of dot plots shows co-expression of IL-17a and IL-22, lower panel shows co-expression of IFN-γ and IL-22. Numbers above each dot plot indicate frequency of positive cells in each quadrant. (B) Frequency of total IL-22-, IL-17a- and IFN-γ-expressing CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 11). (C) Frequency of IL-22 + /IL-17a + and IL-22 + /IL-17a - or IL-22 + /IFN-γ + and IL-22 + /IFN-γ - CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 11). *p

    Article Snippet: For intracellular cytokine staining the following monoclonal antibodies were used: PE-labeled anti-IL-22 (clone 22URTI, eBioscience), PerCP-Cy5.5-labeled anti-IFN-γ (clone B27, BD Biosciences), Alexa Fluor 647-labeled anti-IL-17a (clone N49-653, BD Biosciences), APC-labeled anti-IL-4 (clone 8D4-8, BD Biosciences).

    Techniques: Expressing, Staining

    1,25D diff -DC-supernatant mediated priming of IL-22-producing T cells is dependent on TNF-α IL-6 and IL-23. Supernatants of TLR2/1-stimulated 1,25D diff -DCs were added to naïve CD4 + T cells activated via CD3/CD28-coated beads (as described in Fig 4 ) in the presence or absence of different monoclonal blocking antibodies as indicated. After five days, rIL2 was added to all cultures. On day 12, T cells were restimulated with PMA/Ionomycin for five hours, the last 2.5 hours of culture in the presence of Brefeldin A, in fresh media and intracellular cytokine expression of IL-22, IFN-γ or IL-17a was measured. Cytokine secretion was evaluated after 18–24 hours without further addition of Brefeldin A. (A) Anti-TNF-α, anti-IL-6R-α (5 μg/ml each) or anti-TGF-β (10 μg/ml). T cell-derived IL-22 assessed by ELISA (mean of cytokine levels in ng/ml ± SEM, n = 5). (B) Anti-TNF-α, anti-IL-6R-α (5 μg/ml each) or anti-TGF-β (10 μg/ml). Frequency of total IL-22-expressing CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 5). (C) Anti-IL-1β or anti-IL-23p40 (5 μg/ml each). Frequency of total IL-22-expressing CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 5). (D) Anti-TNF-α, anti-IL-6R-α or anti-IL-23p40 (5 μg/ml each) blocking antibodies alone or in combination. Dot plots from one representative staining of one donor out of five showing the frequency of IL-22-expressing CD4 + T cells against the sideward-scatter (SSC). Numbers in rectangle gate indicate frequency of positive cells. *p

    Journal: PLoS ONE

    Article Title: Vitamin D Regulates Cytokine Patterns Secreted by Dendritic Cells to Promote Differentiation of IL-22-Producing T Cells

    doi: 10.1371/journal.pone.0130395

    Figure Lengend Snippet: 1,25D diff -DC-supernatant mediated priming of IL-22-producing T cells is dependent on TNF-α IL-6 and IL-23. Supernatants of TLR2/1-stimulated 1,25D diff -DCs were added to naïve CD4 + T cells activated via CD3/CD28-coated beads (as described in Fig 4 ) in the presence or absence of different monoclonal blocking antibodies as indicated. After five days, rIL2 was added to all cultures. On day 12, T cells were restimulated with PMA/Ionomycin for five hours, the last 2.5 hours of culture in the presence of Brefeldin A, in fresh media and intracellular cytokine expression of IL-22, IFN-γ or IL-17a was measured. Cytokine secretion was evaluated after 18–24 hours without further addition of Brefeldin A. (A) Anti-TNF-α, anti-IL-6R-α (5 μg/ml each) or anti-TGF-β (10 μg/ml). T cell-derived IL-22 assessed by ELISA (mean of cytokine levels in ng/ml ± SEM, n = 5). (B) Anti-TNF-α, anti-IL-6R-α (5 μg/ml each) or anti-TGF-β (10 μg/ml). Frequency of total IL-22-expressing CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 5). (C) Anti-IL-1β or anti-IL-23p40 (5 μg/ml each). Frequency of total IL-22-expressing CD4 + T cells assessed by intracellular cytokine staining (mean percentage of positive cells ± SEM, n = 5). (D) Anti-TNF-α, anti-IL-6R-α or anti-IL-23p40 (5 μg/ml each) blocking antibodies alone or in combination. Dot plots from one representative staining of one donor out of five showing the frequency of IL-22-expressing CD4 + T cells against the sideward-scatter (SSC). Numbers in rectangle gate indicate frequency of positive cells. *p

    Article Snippet: For intracellular cytokine staining the following monoclonal antibodies were used: PE-labeled anti-IL-22 (clone 22URTI, eBioscience), PerCP-Cy5.5-labeled anti-IFN-γ (clone B27, BD Biosciences), Alexa Fluor 647-labeled anti-IL-17a (clone N49-653, BD Biosciences), APC-labeled anti-IL-4 (clone 8D4-8, BD Biosciences).

    Techniques: Blocking Assay, Expressing, Derivative Assay, Enzyme-linked Immunosorbent Assay, Staining

    The effect of ethyl pyruvate (EP) on Treg in vitro. CD4 + CD25 − T lymphocytes were stimulated for 24 h with a conventional Treg differentiation cocktail (control) and then additionally treated with 125 μM EP. ( A ) Treg (CD4 + CD25 high FoxP3 + ) proportion, at indicated time points after the addition of EP. Representative dot plots for the 72nd h are shown on the right-hand side. ( B ) Carboxyfluorescein succinimidyl ester (CFSE)-based proliferation of Treg (CD4 + CD25 high ) was evaluated by flow cytometry (divided, Div. and undivided, Und. cells, with representative histogram plots shown on the right-hand side) 48 h after EP treatment. Unstimulated cells were stained with CFSE and served as the control for setting the threshold for undivided cells. ( C ) The proportion of Treg expressing perforin, granzyme or CD39, 72 h after EP treatment. Representative dot plots for perforin + Treg are shown on the right-hand side. ( D ) mRNA expression of Tgf-β and Il-10 in cultures treated with or without EP. * p

    Journal: Molecules

    Article Title: Ethyl Pyruvate Promotes Proliferation of Regulatory T Cells by Increasing Glycolysis

    doi: 10.3390/molecules25184112

    Figure Lengend Snippet: The effect of ethyl pyruvate (EP) on Treg in vitro. CD4 + CD25 − T lymphocytes were stimulated for 24 h with a conventional Treg differentiation cocktail (control) and then additionally treated with 125 μM EP. ( A ) Treg (CD4 + CD25 high FoxP3 + ) proportion, at indicated time points after the addition of EP. Representative dot plots for the 72nd h are shown on the right-hand side. ( B ) Carboxyfluorescein succinimidyl ester (CFSE)-based proliferation of Treg (CD4 + CD25 high ) was evaluated by flow cytometry (divided, Div. and undivided, Und. cells, with representative histogram plots shown on the right-hand side) 48 h after EP treatment. Unstimulated cells were stained with CFSE and served as the control for setting the threshold for undivided cells. ( C ) The proportion of Treg expressing perforin, granzyme or CD39, 72 h after EP treatment. Representative dot plots for perforin + Treg are shown on the right-hand side. ( D ) mRNA expression of Tgf-β and Il-10 in cultures treated with or without EP. * p

    Article Snippet: Cells were then fixed in 2% paraformaldehyde, permeabilized with a permeabilization buffer (eBioscience) and stained with anti-mouse IL-10-FITC (rat IgG2b, κ) (eBioscience).

    Techniques: In Vitro, Flow Cytometry, Staining, Expressing