cd279 pd 1 monoclonal antibody  (Thermo Fisher)


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    Name:
    CD279 PD 1 Monoclonal Antibody MIH4
    Description:
    CD279 PD 1 Monoclonal Antibody for Flow
    Catalog Number:
    11-9969-41
    Price:
    None
    Category:
    Antibodies Secondary Detection Reagents
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    Structured Review

    Thermo Fisher cd279 pd 1 monoclonal antibody
    CD279 PD 1 Monoclonal Antibody for Flow
    https://www.bioz.com/result/cd279 pd 1 monoclonal antibody/product/Thermo Fisher
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    cd279 pd 1 monoclonal antibody - by Bioz Stars, 2021-06
    93/100 stars

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    Related Articles

    Staining:

    Article Title: Tumor-infiltrating Tim-3+ T cells proliferate avidly except when PD-1 is co-expressed: Evidence for intracellular cross talk
    Article Snippet: After centrifugation, mononuclear cells were recovered and stored at −80°C until flow cytometry analysis or immediately used for experiments. .. The following anti-human antibodies were used for staining of PBMC or TIL: CD3-Alexa Fluor 700 (BD 557917), CD8+ -PE (BD 555367), CD4+ -PercP/Cy5.5 (BD 560650), FOXP3-PerCP/Cy5.5 (BD 561493), Tim-3-BV421 (Biolegend 345007), CD25-PE-Cy7 (Biolegend 302612), CD4+ -PE-TR (Life Technologies MHCD0417), PD-1-APC (eBioscience 17-9969-42) and CD45RA-PE-Cy7 (eBioscience 25-0458-71), CCR7-FITC (R & D Systems FAB197F) and phospho-S6 (Ser235/236)-Alexa Fluor 488 (Cell Signaling Technology 4803). .. Antibodies to mouse proteins were as follows: Tim-3-PE (R & D, 215008); PD1-APC (Biolegend 109111); CD3-biotin (BD 553060); CD28-biotin (BD 553296).

    Article Title: Mesenchymal stem cells protect against malaria pathogenesis by reprogramming erythropoiesis in the bone marrow
    Article Snippet: Cells were washed and then re-suspended in RPMI 1640 medium supplemented with 10% FBS (Hyclone, USA), 2 mM L-glutamine (Gibco-BRL, Life technologies, USA), and 100 IU/mL penicillin-streptomycin (Gibco-BRL). .. Flow cytometryFor flow cytometric analyses, spleen cells from mice were harvested on the 7th day post-infection, suspended in 50 μl of FACS buffer (PBS, 2% FCS) and surface stained at 4 °C with anti-mouse CD4 (GK1.5, Cat#17-0041-82) -CD8 (53-6.7 Cat#11-0081-82), -CD19 (1D3, Cat#17 0193-82), -CD29 (HM-Bta1-1 Cat#12-0291-82), -CD3 (2C11 Cat#17-0032-82), -Sca-1 (Ly-6A/E Cta#11-5981-82) PD-1 (MIH4 Cat#12-9969-42) were purchased from e-Biosciences and -CD34 (RAM34 Cat#551387 BD Biosciences) Stained cells were acquired with a FACS Fortessa (BD Biosciences) and analysed by Flow Jo (Tree star) or cyflogic (CyFlo Ltd, Finland) software. .. Adoptive transfers of MSCs For adoptive transfer experiments, MSCs were purified by depletion of lineage-differentiated cells such as T lymphocytes, B lymphocytes, macrophages, and DCs (using micro-beads from Miltenyi-Biotec).

    Article Title: Methamphetamine mediates immune dysregulation in a murine model of chronic viral infection
    Article Snippet: After incubation for 5–6 h at 37°C, cells were centrifuged, washed and then stained with PE labeled Pro5® MHC class I Pentamer (Proimmune, 10 μl/test) specific to allele H-2Db and other fluorescence conjugated antibodies to cell surface markers and intracellular cytokines for flow cytometric analysis. .. The following antibodies were used for staining: CD3 APC-Cy7, CD4 AmCyan, CD8 Pacific Blue, LCMV Pentamer PE, CD279 (PD1) PECy7, CD25 PerCpCy5.5, CD69 APC, CD27 PeCy7, CXCR3 PerCpCy5.5, and EGFR FITC (antibodies were from eBiosciences, San Diego, and BD Biosciences, CA, USA). .. Stained cells were then acquired on a FACS Canto II (BD Biosciences) using FACS Diva software (BD Biosciences, V 6.1.3).

    Mouse Assay:

    Article Title: Mesenchymal stem cells protect against malaria pathogenesis by reprogramming erythropoiesis in the bone marrow
    Article Snippet: Cells were washed and then re-suspended in RPMI 1640 medium supplemented with 10% FBS (Hyclone, USA), 2 mM L-glutamine (Gibco-BRL, Life technologies, USA), and 100 IU/mL penicillin-streptomycin (Gibco-BRL). .. Flow cytometryFor flow cytometric analyses, spleen cells from mice were harvested on the 7th day post-infection, suspended in 50 μl of FACS buffer (PBS, 2% FCS) and surface stained at 4 °C with anti-mouse CD4 (GK1.5, Cat#17-0041-82) -CD8 (53-6.7 Cat#11-0081-82), -CD19 (1D3, Cat#17 0193-82), -CD29 (HM-Bta1-1 Cat#12-0291-82), -CD3 (2C11 Cat#17-0032-82), -Sca-1 (Ly-6A/E Cta#11-5981-82) PD-1 (MIH4 Cat#12-9969-42) were purchased from e-Biosciences and -CD34 (RAM34 Cat#551387 BD Biosciences) Stained cells were acquired with a FACS Fortessa (BD Biosciences) and analysed by Flow Jo (Tree star) or cyflogic (CyFlo Ltd, Finland) software. .. Adoptive transfers of MSCs For adoptive transfer experiments, MSCs were purified by depletion of lineage-differentiated cells such as T lymphocytes, B lymphocytes, macrophages, and DCs (using micro-beads from Miltenyi-Biotec).

    FACS:

    Article Title: Mesenchymal stem cells protect against malaria pathogenesis by reprogramming erythropoiesis in the bone marrow
    Article Snippet: Cells were washed and then re-suspended in RPMI 1640 medium supplemented with 10% FBS (Hyclone, USA), 2 mM L-glutamine (Gibco-BRL, Life technologies, USA), and 100 IU/mL penicillin-streptomycin (Gibco-BRL). .. Flow cytometryFor flow cytometric analyses, spleen cells from mice were harvested on the 7th day post-infection, suspended in 50 μl of FACS buffer (PBS, 2% FCS) and surface stained at 4 °C with anti-mouse CD4 (GK1.5, Cat#17-0041-82) -CD8 (53-6.7 Cat#11-0081-82), -CD19 (1D3, Cat#17 0193-82), -CD29 (HM-Bta1-1 Cat#12-0291-82), -CD3 (2C11 Cat#17-0032-82), -Sca-1 (Ly-6A/E Cta#11-5981-82) PD-1 (MIH4 Cat#12-9969-42) were purchased from e-Biosciences and -CD34 (RAM34 Cat#551387 BD Biosciences) Stained cells were acquired with a FACS Fortessa (BD Biosciences) and analysed by Flow Jo (Tree star) or cyflogic (CyFlo Ltd, Finland) software. .. Adoptive transfers of MSCs For adoptive transfer experiments, MSCs were purified by depletion of lineage-differentiated cells such as T lymphocytes, B lymphocytes, macrophages, and DCs (using micro-beads from Miltenyi-Biotec).

    Software:

    Article Title: Mesenchymal stem cells protect against malaria pathogenesis by reprogramming erythropoiesis in the bone marrow
    Article Snippet: Cells were washed and then re-suspended in RPMI 1640 medium supplemented with 10% FBS (Hyclone, USA), 2 mM L-glutamine (Gibco-BRL, Life technologies, USA), and 100 IU/mL penicillin-streptomycin (Gibco-BRL). .. Flow cytometryFor flow cytometric analyses, spleen cells from mice were harvested on the 7th day post-infection, suspended in 50 μl of FACS buffer (PBS, 2% FCS) and surface stained at 4 °C with anti-mouse CD4 (GK1.5, Cat#17-0041-82) -CD8 (53-6.7 Cat#11-0081-82), -CD19 (1D3, Cat#17 0193-82), -CD29 (HM-Bta1-1 Cat#12-0291-82), -CD3 (2C11 Cat#17-0032-82), -Sca-1 (Ly-6A/E Cta#11-5981-82) PD-1 (MIH4 Cat#12-9969-42) were purchased from e-Biosciences and -CD34 (RAM34 Cat#551387 BD Biosciences) Stained cells were acquired with a FACS Fortessa (BD Biosciences) and analysed by Flow Jo (Tree star) or cyflogic (CyFlo Ltd, Finland) software. .. Adoptive transfers of MSCs For adoptive transfer experiments, MSCs were purified by depletion of lineage-differentiated cells such as T lymphocytes, B lymphocytes, macrophages, and DCs (using micro-beads from Miltenyi-Biotec).

    Chloramphenicol Acetyltransferase Assay:

    Article Title: Immunogenic neoantigens derived from gene fusions stimulate T cell responses
    Article Snippet: For immunohistochemical staining of MSK-HN1 tumor samples, the antibodies are listed with the supplier, catalogue number, and dilution: anti-CD3 (Leica cat# NCL-L-CD3–565, 1:100), anti-CD8 (Ventana, cat#790–4460, no dilution), anti-CD45 (Ventana cat# 760–2505, no diution), and anti-PD-L1 (Cell Signaling ca# 13684, 1:800). .. For cell-based assays, the antibodies used are listed with the supplier, catalogue number, and dilution: PE-anti-HLA-A2 clone BB7.2 (BD cat # 343306, 1:500), Alexa Fluor 594 anti-HLA-A clone EP1395Y (Abcam cat# 207872, 1:1,000), polyclonal anti-HLA-B (Invitrogen cat# PA5–35345, 1:1,000), anti-HLA-C clone DT9 (Biolegend cat# 373302, 1:1,000), PE-Cy5.5-F(ab’)2-goat anti-mouse-IgG H+L (Invitrogen cat# M35018, 1:200), PE-F(ab’)2-goat anti-rabbit IgG H+L (Invitrogen cat31864, 1:200), Alexa Fluor 405 anti-CD3e Clone UCHT1 (Thermo cat# CD0326, 1:100), PerCP/Cy5.5 anti-CD4 Clone A161A1 (Biolegend cat# 357414, 1:100), APC-H7 anti-CD8 Clone SKI (BD cat# 560179, 1:100), APC anti-PD-1 Clone M1H4 (eBioscience cat# 17–9969–42, 1:100), PE/Dazzle 594 anti-CD137 Clone 4B4–1 (Biolegend cat# 309825, 1:100), PE anti-CD40L Clone 24–31 (Biolegend cat# 310805, 1:100), Aqua Live/Dead (cat# L34966, 1:1,000), PE anti-active caspase-3 (BD cat# 550821, 1:50), anti-c-Myb Clone D2R4Y (Cell Signaling cat# 12319, 1:1,000), anti-DEK Clone E1L3V (Cell Signaling cat# 13962, 1:1,000), anti-HLA-ABC Monoclonal Antibody Clone W6/32 (Memorial Sloan Kettering Cancer Center antibody and bioresource core facility). .. Histology and immunohistochemistry Histopathologic analyses were conducted by subspecialty pathologists (R.G. and N.K.).

    Flow Cytometry:

    Article Title: Diacylglycerol Kinase ζ Limits Cytokine-dependent Expansion of CD8+ T Cells with Broad Antitumor Capacity
    Article Snippet: Cells were divided every 2–3 days and used for flow cytometry analysis, real-time RT-PCR, degranulation or cytolytic assays on days 6–8. .. 2.4 Flow Cytometry and Antibodies Antibodies used were anti-CD8-PeCy7, -CD3-PERCP Cy5.5, -CD44-PeCy5, -CD25-PeCy7, -CD335-APC (NKP46) (BioLegend), -CD8-eFluor450, CD3-Pecy7, -CD122-FITC, -IL15-receptorα-PE, -CD279-eFluor780 (PD-1), -CD314-PE (NKG2D) (eBioscience), -CD4-PE, -CD3-FITC (Beckman Coulter), and -CD69-PE (Pharmingen). .. Samples were collected on a LSRII (BD Biosciences) or a Cytomics FC 500 (Beckman Coulter) and analyzed with FlowJo software (Tree Star, Ashland, OR).

    Article Title: Monoclonal antibodies specific to human Δ42PD1: A novel immunoregulator potentially involved in HIV-1 and tumor pathogenesis
    Article Snippet: After washing, cells were resuspended in 0.4 ml PBS and then subjected to acquisition with FACSCalibur or FACSAriaIII Flow Cytometer (BD Biosciences), and data were analyzed with FlowJo software. .. Commercial antibodies used in flow cytometry include Alexa Fluor 647 Goat Anti-Mouse IgG (H+ L)(Cat. No. , Life Technologies), Goat anti-Human PD1 polyclonal antibody (Cat. No. AF1086, R & D Systems), Mouse anti-Human PD1 (clone MIH4, Cat. No. 14–9969–82, eBioscience), Mouse IgG1 (clone P3.6.2.8.1, Cat. No. 16–4714–85, eBioscience), Mouse IgG2b (clone eBMG2b, Cat. No. 16–4732–85, eBioscience), Mouse Anti-Human PD1 antibody (clone NAT105, Cat. No. ab52587, Abcam), Alexa Fluor 568 Goat Anti-Mouse IgG1 (Cat. No. A-21124, Life Technologies); Alexa Fluor 488 Goat Anti-Mouse IgG1 (Cat. No. A-21121, Life Technologies); Alexa Fluor 488 Goat Anti-Mouse IgG2b (Cat. No. A-21141, Life Technologies). .. Antibody competitive assay was conducted by using equal amount of soluble mAbs CH101 and CH34 to block the binding of 0.25 μg of pre-labeled Alexa Fluor 467-CH101 to 293T- Δ42PD1 cells.

    Cytometry:

    Article Title: Diacylglycerol Kinase ζ Limits Cytokine-dependent Expansion of CD8+ T Cells with Broad Antitumor Capacity
    Article Snippet: Cells were divided every 2–3 days and used for flow cytometry analysis, real-time RT-PCR, degranulation or cytolytic assays on days 6–8. .. 2.4 Flow Cytometry and Antibodies Antibodies used were anti-CD8-PeCy7, -CD3-PERCP Cy5.5, -CD44-PeCy5, -CD25-PeCy7, -CD335-APC (NKP46) (BioLegend), -CD8-eFluor450, CD3-Pecy7, -CD122-FITC, -IL15-receptorα-PE, -CD279-eFluor780 (PD-1), -CD314-PE (NKG2D) (eBioscience), -CD4-PE, -CD3-FITC (Beckman Coulter), and -CD69-PE (Pharmingen). .. Samples were collected on a LSRII (BD Biosciences) or a Cytomics FC 500 (Beckman Coulter) and analyzed with FlowJo software (Tree Star, Ashland, OR).

    Article Title: Monoclonal antibodies specific to human Δ42PD1: A novel immunoregulator potentially involved in HIV-1 and tumor pathogenesis
    Article Snippet: After washing, cells were resuspended in 0.4 ml PBS and then subjected to acquisition with FACSCalibur or FACSAriaIII Flow Cytometer (BD Biosciences), and data were analyzed with FlowJo software. .. Commercial antibodies used in flow cytometry include Alexa Fluor 647 Goat Anti-Mouse IgG (H+ L)(Cat. No. , Life Technologies), Goat anti-Human PD1 polyclonal antibody (Cat. No. AF1086, R & D Systems), Mouse anti-Human PD1 (clone MIH4, Cat. No. 14–9969–82, eBioscience), Mouse IgG1 (clone P3.6.2.8.1, Cat. No. 16–4714–85, eBioscience), Mouse IgG2b (clone eBMG2b, Cat. No. 16–4732–85, eBioscience), Mouse Anti-Human PD1 antibody (clone NAT105, Cat. No. ab52587, Abcam), Alexa Fluor 568 Goat Anti-Mouse IgG1 (Cat. No. A-21124, Life Technologies); Alexa Fluor 488 Goat Anti-Mouse IgG1 (Cat. No. A-21121, Life Technologies); Alexa Fluor 488 Goat Anti-Mouse IgG2b (Cat. No. A-21141, Life Technologies). .. Antibody competitive assay was conducted by using equal amount of soluble mAbs CH101 and CH34 to block the binding of 0.25 μg of pre-labeled Alexa Fluor 467-CH101 to 293T- Δ42PD1 cells.

    Labeling:

    Article Title: Development and Fit-for-Purpose Validation of a Soluble Human Programmed Death-1 Protein Assay
    Article Snippet: Recombinant human PDL-1 and PDL-2 proteins were purchased from R & D Systems (Minneapolis, MN, USA). .. Anti-PD-1 antibody MIH4 (eBiosciences, San Diego, CA, USA) was labeled with Biotin, at 1:20 (antibody: biotin) molar ratio using a EZ-Link NHS-PEG4-Biotin kit (Thermo Fisher, Waltham, MA, USA). .. An anti-PD-1 antibody, AF1086 from R & D Systems, were labeled with ruthenium, at 1:12 (antibody:ruthenium) molar ratio, using the MSD SULFO-TAG NHS-Ester kit (Meso Scale Discovery, Rockville, MD, USA).

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    Thermo Fisher mouse anti human pd 1 antibody
    <t>PD-1</t> blockade decreases Bim expression induced by PD-L1 in human CD8 + T cells.
    Mouse Anti Human Pd 1 Antibody, 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/mouse anti human pd 1 antibody/product/Thermo Fisher
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    Thermo Fisher pe conjugated anti pd 1
    <t>PD-1</t> deletion alters Treg homeostasis during IL-2 therapy. C57BL/6 PD-1 −/− or C57BL/6 WT mice were treated with control vehicle or IL-2 at 5000 IU once per day for 4 weeks. (A) Effect of IL-2 therapy on frequency of Tregs during treatment. Increase of percentage of Tregs during therapy from the baseline level of each group. (B-H) Spleen cells were analyzed after 1 week of IL-2 therapy. (B) pSTAT5 expression in Tregs. (C) Representative flow cytometry histograms detecting Ki-67 + proliferating Tregs. Percentage of Ki-67 + Tregs is shown for each histogram. (D) Percentage of Ki-67 + proliferating Tregs in PD-1 −/− and PD-1 WT mice. (E) Representative histogram identifying CD4 Tregs in PD-1 −/− and PD-1 WT mice receiving control vehicle or IL-2. (F) Frequency of CD4 + CD25 + Foxp3 + Tregs in spleen (percentage of CD4 T cells). (G) Number of CD4 + CD25 + Foxp3 + Tregs in spleen. (H-L) Spleen cells were analyzed after 2 weeks IL-2 therapy given once per day. (H) Representative histograms identify CD44 low CD62L high naïve, CD44 high CD62L high central-memory, and CD44 high CD62L low effector-memory Treg subsets after IL-2 therapy. (I) Percentage of each Treg subset after IL-2 therapy. (J) Percentage of annexin-V + apoptotic cells in each Treg subset after IL-2 therapy. (K) Mean fluorescence intensity (MFI) for the ratio of Bcl-2 expression in Tregs of IL-2–treated mice:Bcl-2 expression in Tregs of control vehicle–treated mice. (L) MFI for the ratio of Fas expression in Tregs of IL-2–treated mice:Fas expression in Tregs of control vehicle–treated mice with 4 mice per group per experiment. Data are representative of (A) 2 or (B-L) 3 independent experiments and expressed as means +/– SEM. * P
    Pe Conjugated Anti Pd 1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher pd1
    TBK1 is required for T FH cell differentiation. ( a , c , e ) Flow cytometry of cells from host B6 mice 7 d after adoptive transfer of SMARTA CD4 + T cells transduced with shRNA targeting the Tbk1 (sh Tbk1 -1 and sh Tbk1 -2), Icos or control genes and infected with LCMV Armstrong strain. Numbers adjacent to outlined areas indicate percent CXCR5 + SLAM lo T FH cells ( a ), CXCR5 + <t>PD1</t> hi GC T FH cells ( c ), or CXCR5 + GL7 hi GC T FH cells ( e ). Cumulative data for a ( b ), c ( d ) or e ( f ) from three independent experiments. ( g ) Flow cytometry of cells from host CD4-Cre × Bcl6 fl/fl mice 10 d after adoptive transfer of Icos –/– SMARTA CD4 + T cells transduced as in ( a ), and were immunized with KLH-gp61 suspended in AddaVax. Numbers adjacent to outlined areas indicate percent CD95 + PNA + GC B cells. Cumulative data for g ( h ) from two independent experiments. ( i ) Quantification of anti-KLH-gp61 IgG from sera of immunized mice analyzed with ELISA and presented as absorbance at 450 nm. The endpoint titer ( j ) and area under curve (AUC; k ) were calculated. Each data point represents a single mouse. Shown are mean ± SEM; ANOVA with post-hoc Tukey's corrections. * P
    Pd1, 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
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    Thermo Fisher anti pdl1
    Activation of PI3K/Akt signalling regulates neuronal FoxA1 and <t>PDL1.</t> ( a ) Real-time PCR analysed fold changes of Pi3k and Akt mRNA in CGN. Graphs are mean±s.e.m. from three independent experiments. Two-way analysis of variance (ANOVA) test was used, * P
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    Image Search Results


    PD-1 blockade decreases Bim expression induced by PD-L1 in human CD8 + T cells.

    Journal: JCI Insight

    Article Title: T cell Bim levels reflect responses to anti–PD-1 cancer therapy

    doi: 10.1172/jci.insight.86014

    Figure Lengend Snippet: PD-1 blockade decreases Bim expression induced by PD-L1 in human CD8 + T cells.

    Article Snippet: To block PD-1/PD-L1 interaction-induced Bim upregulation, preactivated CD8+ T cells were incubated with 10 μg/ml of mouse anti-human PD-1 antibody (clone MIH4, eBioscience, special order for functional grade-purified version) or humanized antibody to PD-1 (Keytruda or Opdivo, Mayo Clinic Pharmacy) for 20 minutes before culture with PD-L1 fusion protein.

    Techniques: Expressing

    Bim upregulation is associated with PD-1 expression in melanoma patients.

    Journal: JCI Insight

    Article Title: T cell Bim levels reflect responses to anti–PD-1 cancer therapy

    doi: 10.1172/jci.insight.86014

    Figure Lengend Snippet: Bim upregulation is associated with PD-1 expression in melanoma patients.

    Article Snippet: To block PD-1/PD-L1 interaction-induced Bim upregulation, preactivated CD8+ T cells were incubated with 10 μg/ml of mouse anti-human PD-1 antibody (clone MIH4, eBioscience, special order for functional grade-purified version) or humanized antibody to PD-1 (Keytruda or Opdivo, Mayo Clinic Pharmacy) for 20 minutes before culture with PD-L1 fusion protein.

    Techniques: Expressing

    Changes of Bim levels predict responses to anti–PD-1 therapy in melanoma patients.

    Journal: JCI Insight

    Article Title: T cell Bim levels reflect responses to anti–PD-1 cancer therapy

    doi: 10.1172/jci.insight.86014

    Figure Lengend Snippet: Changes of Bim levels predict responses to anti–PD-1 therapy in melanoma patients.

    Article Snippet: To block PD-1/PD-L1 interaction-induced Bim upregulation, preactivated CD8+ T cells were incubated with 10 μg/ml of mouse anti-human PD-1 antibody (clone MIH4, eBioscience, special order for functional grade-purified version) or humanized antibody to PD-1 (Keytruda or Opdivo, Mayo Clinic Pharmacy) for 20 minutes before culture with PD-L1 fusion protein.

    Techniques:

    Bim upregulation in tumor-reactive CD8 + T cells is dependent on PD-1 and PD-L1 interaction.

    Journal: JCI Insight

    Article Title: T cell Bim levels reflect responses to anti–PD-1 cancer therapy

    doi: 10.1172/jci.insight.86014

    Figure Lengend Snippet: Bim upregulation in tumor-reactive CD8 + T cells is dependent on PD-1 and PD-L1 interaction.

    Article Snippet: To block PD-1/PD-L1 interaction-induced Bim upregulation, preactivated CD8+ T cells were incubated with 10 μg/ml of mouse anti-human PD-1 antibody (clone MIH4, eBioscience, special order for functional grade-purified version) or humanized antibody to PD-1 (Keytruda or Opdivo, Mayo Clinic Pharmacy) for 20 minutes before culture with PD-L1 fusion protein.

    Techniques:

    PD-1 deletion alters Treg homeostasis during IL-2 therapy. C57BL/6 PD-1 −/− or C57BL/6 WT mice were treated with control vehicle or IL-2 at 5000 IU once per day for 4 weeks. (A) Effect of IL-2 therapy on frequency of Tregs during treatment. Increase of percentage of Tregs during therapy from the baseline level of each group. (B-H) Spleen cells were analyzed after 1 week of IL-2 therapy. (B) pSTAT5 expression in Tregs. (C) Representative flow cytometry histograms detecting Ki-67 + proliferating Tregs. Percentage of Ki-67 + Tregs is shown for each histogram. (D) Percentage of Ki-67 + proliferating Tregs in PD-1 −/− and PD-1 WT mice. (E) Representative histogram identifying CD4 Tregs in PD-1 −/− and PD-1 WT mice receiving control vehicle or IL-2. (F) Frequency of CD4 + CD25 + Foxp3 + Tregs in spleen (percentage of CD4 T cells). (G) Number of CD4 + CD25 + Foxp3 + Tregs in spleen. (H-L) Spleen cells were analyzed after 2 weeks IL-2 therapy given once per day. (H) Representative histograms identify CD44 low CD62L high naïve, CD44 high CD62L high central-memory, and CD44 high CD62L low effector-memory Treg subsets after IL-2 therapy. (I) Percentage of each Treg subset after IL-2 therapy. (J) Percentage of annexin-V + apoptotic cells in each Treg subset after IL-2 therapy. (K) Mean fluorescence intensity (MFI) for the ratio of Bcl-2 expression in Tregs of IL-2–treated mice:Bcl-2 expression in Tregs of control vehicle–treated mice. (L) MFI for the ratio of Fas expression in Tregs of IL-2–treated mice:Fas expression in Tregs of control vehicle–treated mice with 4 mice per group per experiment. Data are representative of (A) 2 or (B-L) 3 independent experiments and expressed as means +/– SEM. * P

    Journal: Blood

    Article Title: PD-1 modulates regulatory T-cell homeostasis during low-dose interleukin-2 therapy

    doi: 10.1182/blood-2016-09-741629

    Figure Lengend Snippet: PD-1 deletion alters Treg homeostasis during IL-2 therapy. C57BL/6 PD-1 −/− or C57BL/6 WT mice were treated with control vehicle or IL-2 at 5000 IU once per day for 4 weeks. (A) Effect of IL-2 therapy on frequency of Tregs during treatment. Increase of percentage of Tregs during therapy from the baseline level of each group. (B-H) Spleen cells were analyzed after 1 week of IL-2 therapy. (B) pSTAT5 expression in Tregs. (C) Representative flow cytometry histograms detecting Ki-67 + proliferating Tregs. Percentage of Ki-67 + Tregs is shown for each histogram. (D) Percentage of Ki-67 + proliferating Tregs in PD-1 −/− and PD-1 WT mice. (E) Representative histogram identifying CD4 Tregs in PD-1 −/− and PD-1 WT mice receiving control vehicle or IL-2. (F) Frequency of CD4 + CD25 + Foxp3 + Tregs in spleen (percentage of CD4 T cells). (G) Number of CD4 + CD25 + Foxp3 + Tregs in spleen. (H-L) Spleen cells were analyzed after 2 weeks IL-2 therapy given once per day. (H) Representative histograms identify CD44 low CD62L high naïve, CD44 high CD62L high central-memory, and CD44 high CD62L low effector-memory Treg subsets after IL-2 therapy. (I) Percentage of each Treg subset after IL-2 therapy. (J) Percentage of annexin-V + apoptotic cells in each Treg subset after IL-2 therapy. (K) Mean fluorescence intensity (MFI) for the ratio of Bcl-2 expression in Tregs of IL-2–treated mice:Bcl-2 expression in Tregs of control vehicle–treated mice. (L) MFI for the ratio of Fas expression in Tregs of IL-2–treated mice:Fas expression in Tregs of control vehicle–treated mice with 4 mice per group per experiment. Data are representative of (A) 2 or (B-L) 3 independent experiments and expressed as means +/– SEM. * P

    Article Snippet: Peripheral blood mononuclear cells (PBMCs) were incubated with the following directly conjugated monoclonal antibodies for 20 minutes at 4°C: Pacific Blue–conjugated anti - CD4 (RPA-T4; BD Biosciences), PE-Cy7–conjugated anti - CD25 (M-A251; BD Biosciences), FITC-conjugated anti - CD45RA (ALB11; Beckman Coulter), PE-conjugated anti-PD-1 (eBioJ105; eBioscience), and APC-Fluor750–conjugated anti - CD127 (eBioRDR5; eBioscience).

    Techniques: Mouse Assay, Expressing, Flow Cytometry, Cytometry, Fluorescence

    Phenotypic changes in murine Tregs after IL-2 therapy. Wild-type C57BL/6 mice received control vehicle or 5000 IU human recombinant IL-2 subcutaneously once per day for 14 days and spleen cells were analyzed on day 15. (A) Representative panels identify CD44 low CD62L high naïve (N), CD44 high CD62L high central-memory (CM), and CD44 high CD62L low effector-memory (EM) subsets within CD8 T cells, Tcons, and Tregs. (B) Percentage of each subset in CD8 T cells, Tcons, and Tregs after in vivo treatment with control vehicle or IL-2. (C) Representative flow cytometry histograms identifying PD-1 + cells in each T-cell subset after treatment with control vehicle or IL-2. (D) Percentage of PD-1 + cells in CD8 T cells, Tcons, and Tregs after treatment with control vehicle or IL-2. (E) Representative flow cytometry histograms detecting expression of CTLA-4, LAG-3, Tim-3, and PD-1 on CD8 T cells, Tcons, and Tregs with 4 mice per group per experiment. Data are representative of 3 independent experiments and expressed as means +/– SEM. *** P

    Journal: Blood

    Article Title: PD-1 modulates regulatory T-cell homeostasis during low-dose interleukin-2 therapy

    doi: 10.1182/blood-2016-09-741629

    Figure Lengend Snippet: Phenotypic changes in murine Tregs after IL-2 therapy. Wild-type C57BL/6 mice received control vehicle or 5000 IU human recombinant IL-2 subcutaneously once per day for 14 days and spleen cells were analyzed on day 15. (A) Representative panels identify CD44 low CD62L high naïve (N), CD44 high CD62L high central-memory (CM), and CD44 high CD62L low effector-memory (EM) subsets within CD8 T cells, Tcons, and Tregs. (B) Percentage of each subset in CD8 T cells, Tcons, and Tregs after in vivo treatment with control vehicle or IL-2. (C) Representative flow cytometry histograms identifying PD-1 + cells in each T-cell subset after treatment with control vehicle or IL-2. (D) Percentage of PD-1 + cells in CD8 T cells, Tcons, and Tregs after treatment with control vehicle or IL-2. (E) Representative flow cytometry histograms detecting expression of CTLA-4, LAG-3, Tim-3, and PD-1 on CD8 T cells, Tcons, and Tregs with 4 mice per group per experiment. Data are representative of 3 independent experiments and expressed as means +/– SEM. *** P

    Article Snippet: Peripheral blood mononuclear cells (PBMCs) were incubated with the following directly conjugated monoclonal antibodies for 20 minutes at 4°C: Pacific Blue–conjugated anti - CD4 (RPA-T4; BD Biosciences), PE-Cy7–conjugated anti - CD25 (M-A251; BD Biosciences), FITC-conjugated anti - CD45RA (ALB11; Beckman Coulter), PE-conjugated anti-PD-1 (eBioJ105; eBioscience), and APC-Fluor750–conjugated anti - CD127 (eBioRDR5; eBioscience).

    Techniques: Mouse Assay, Recombinant, In Vivo, Flow Cytometry, Cytometry, Expressing

    Selective increase of PD-1 expression on CD45RA – activated-memory Tregs in patients with cGVHD receiving low-dose IL-2. (A) Representative flow cytometry histograms used to define CD4 T-cell subsets. (B) Median percentages of PD-1 + cells in Tcon and Treg subsets during IL-2 therapy; CD45RA – activated-memory Tregs vs baseline using Wilcoxon signed rank test. * P

    Journal: Blood

    Article Title: PD-1 modulates regulatory T-cell homeostasis during low-dose interleukin-2 therapy

    doi: 10.1182/blood-2016-09-741629

    Figure Lengend Snippet: Selective increase of PD-1 expression on CD45RA – activated-memory Tregs in patients with cGVHD receiving low-dose IL-2. (A) Representative flow cytometry histograms used to define CD4 T-cell subsets. (B) Median percentages of PD-1 + cells in Tcon and Treg subsets during IL-2 therapy; CD45RA – activated-memory Tregs vs baseline using Wilcoxon signed rank test. * P

    Article Snippet: Peripheral blood mononuclear cells (PBMCs) were incubated with the following directly conjugated monoclonal antibodies for 20 minutes at 4°C: Pacific Blue–conjugated anti - CD4 (RPA-T4; BD Biosciences), PE-Cy7–conjugated anti - CD25 (M-A251; BD Biosciences), FITC-conjugated anti - CD45RA (ALB11; Beckman Coulter), PE-conjugated anti-PD-1 (eBioJ105; eBioscience), and APC-Fluor750–conjugated anti - CD127 (eBioRDR5; eBioscience).

    Techniques: Expressing, Flow Cytometry, Cytometry

    Effects of combined IL-2 therapy and PD-1 blockade on Treg expansion in vivo. Wild-type C57BL/6 mice received vehicle (plus isotype antibody), IL-2 (plus isotype antibody), anti-PD-1 (aPD-1) antibody (plus vehicle control), or IL-2 plus anti-PD-1 antibody. Anti-PD-1 antibody (250 μg) was administrated intraperitoneally twice per week for a total of 4 injections beginning on the first day of IL-2 treatment. IL-2–treated groups received IL-2 at 5000 IU once per day for 14 days. Peripheral blood cells were collected and analyzed at days 0, 4, 8, 11, and 15. (A) Increase of the percentage of Ki-67 + proliferating Tregs from the baseline level of each group during therapy. (B) Increase of the percentage of Tregs during therapy from the baseline level of each group with 4 mice per group per experiment. Data are representative of 2 independent experiments and expressed as means +/– SEM. * P

    Journal: Blood

    Article Title: PD-1 modulates regulatory T-cell homeostasis during low-dose interleukin-2 therapy

    doi: 10.1182/blood-2016-09-741629

    Figure Lengend Snippet: Effects of combined IL-2 therapy and PD-1 blockade on Treg expansion in vivo. Wild-type C57BL/6 mice received vehicle (plus isotype antibody), IL-2 (plus isotype antibody), anti-PD-1 (aPD-1) antibody (plus vehicle control), or IL-2 plus anti-PD-1 antibody. Anti-PD-1 antibody (250 μg) was administrated intraperitoneally twice per week for a total of 4 injections beginning on the first day of IL-2 treatment. IL-2–treated groups received IL-2 at 5000 IU once per day for 14 days. Peripheral blood cells were collected and analyzed at days 0, 4, 8, 11, and 15. (A) Increase of the percentage of Ki-67 + proliferating Tregs from the baseline level of each group during therapy. (B) Increase of the percentage of Tregs during therapy from the baseline level of each group with 4 mice per group per experiment. Data are representative of 2 independent experiments and expressed as means +/– SEM. * P

    Article Snippet: Peripheral blood mononuclear cells (PBMCs) were incubated with the following directly conjugated monoclonal antibodies for 20 minutes at 4°C: Pacific Blue–conjugated anti - CD4 (RPA-T4; BD Biosciences), PE-Cy7–conjugated anti - CD25 (M-A251; BD Biosciences), FITC-conjugated anti - CD45RA (ALB11; Beckman Coulter), PE-conjugated anti-PD-1 (eBioJ105; eBioscience), and APC-Fluor750–conjugated anti - CD127 (eBioRDR5; eBioscience).

    Techniques: In Vivo, Mouse Assay

    Comparison of PD-1 expression in clinical responders and nonresponders during low-dose IL-2 therapy. ( A ) Scatter plot of the percentage of PD-1 + CD45RA – Tregs and Tcons in nonresponders and responders at week 2 during IL-2 therapy. ( B ) Ratio of Treg percentage of PD-1 + :Tcon percentage of PD-1 in nonresponders and responders before and 2 weeks after starting IL-2 therapy. The ratio is significantly increased in clinical responders 2 weeks after IL-2 administration ( P = .03, Wilcoxon signed rank test). Median values are shown in green.

    Journal: Blood

    Article Title: PD-1 modulates regulatory T-cell homeostasis during low-dose interleukin-2 therapy

    doi: 10.1182/blood-2016-09-741629

    Figure Lengend Snippet: Comparison of PD-1 expression in clinical responders and nonresponders during low-dose IL-2 therapy. ( A ) Scatter plot of the percentage of PD-1 + CD45RA – Tregs and Tcons in nonresponders and responders at week 2 during IL-2 therapy. ( B ) Ratio of Treg percentage of PD-1 + :Tcon percentage of PD-1 in nonresponders and responders before and 2 weeks after starting IL-2 therapy. The ratio is significantly increased in clinical responders 2 weeks after IL-2 administration ( P = .03, Wilcoxon signed rank test). Median values are shown in green.

    Article Snippet: Peripheral blood mononuclear cells (PBMCs) were incubated with the following directly conjugated monoclonal antibodies for 20 minutes at 4°C: Pacific Blue–conjugated anti - CD4 (RPA-T4; BD Biosciences), PE-Cy7–conjugated anti - CD25 (M-A251; BD Biosciences), FITC-conjugated anti - CD45RA (ALB11; Beckman Coulter), PE-conjugated anti-PD-1 (eBioJ105; eBioscience), and APC-Fluor750–conjugated anti - CD127 (eBioRDR5; eBioscience).

    Techniques: Expressing

    PD-1 blockade enhances IL-2–induced proliferation of expanded human Tregs and promotes apoptosis. Purified CD45RA + naïve Tregs and Tcons labeled with carboxyfluorescein succinimidyl ester (CFSE) were stimulated with IL-2, anti-CD3/28 antibody, or anti-PD-L1 antibody for 4 days. Representative flow cytometry histograms were used to quantify CFSE dilution and (A) identify PD-1 + cells and (B) identify annexin-V + cells. (C) Percentage of annexin-V + apoptotic cells within Tcon and Treg populations stimulated with IL-2, anti-CD3/28 antibody, or anti-PD-L1 antibody for 4 days. Data are obtained from 1 experiment.

    Journal: Blood

    Article Title: PD-1 modulates regulatory T-cell homeostasis during low-dose interleukin-2 therapy

    doi: 10.1182/blood-2016-09-741629

    Figure Lengend Snippet: PD-1 blockade enhances IL-2–induced proliferation of expanded human Tregs and promotes apoptosis. Purified CD45RA + naïve Tregs and Tcons labeled with carboxyfluorescein succinimidyl ester (CFSE) were stimulated with IL-2, anti-CD3/28 antibody, or anti-PD-L1 antibody for 4 days. Representative flow cytometry histograms were used to quantify CFSE dilution and (A) identify PD-1 + cells and (B) identify annexin-V + cells. (C) Percentage of annexin-V + apoptotic cells within Tcon and Treg populations stimulated with IL-2, anti-CD3/28 antibody, or anti-PD-L1 antibody for 4 days. Data are obtained from 1 experiment.

    Article Snippet: Peripheral blood mononuclear cells (PBMCs) were incubated with the following directly conjugated monoclonal antibodies for 20 minutes at 4°C: Pacific Blue–conjugated anti - CD4 (RPA-T4; BD Biosciences), PE-Cy7–conjugated anti - CD25 (M-A251; BD Biosciences), FITC-conjugated anti - CD45RA (ALB11; Beckman Coulter), PE-conjugated anti-PD-1 (eBioJ105; eBioscience), and APC-Fluor750–conjugated anti - CD127 (eBioRDR5; eBioscience).

    Techniques: Purification, Labeling, Flow Cytometry, Cytometry

    TBK1 is required for T FH cell differentiation. ( a , c , e ) Flow cytometry of cells from host B6 mice 7 d after adoptive transfer of SMARTA CD4 + T cells transduced with shRNA targeting the Tbk1 (sh Tbk1 -1 and sh Tbk1 -2), Icos or control genes and infected with LCMV Armstrong strain. Numbers adjacent to outlined areas indicate percent CXCR5 + SLAM lo T FH cells ( a ), CXCR5 + PD1 hi GC T FH cells ( c ), or CXCR5 + GL7 hi GC T FH cells ( e ). Cumulative data for a ( b ), c ( d ) or e ( f ) from three independent experiments. ( g ) Flow cytometry of cells from host CD4-Cre × Bcl6 fl/fl mice 10 d after adoptive transfer of Icos –/– SMARTA CD4 + T cells transduced as in ( a ), and were immunized with KLH-gp61 suspended in AddaVax. Numbers adjacent to outlined areas indicate percent CD95 + PNA + GC B cells. Cumulative data for g ( h ) from two independent experiments. ( i ) Quantification of anti-KLH-gp61 IgG from sera of immunized mice analyzed with ELISA and presented as absorbance at 450 nm. The endpoint titer ( j ) and area under curve (AUC; k ) were calculated. Each data point represents a single mouse. Shown are mean ± SEM; ANOVA with post-hoc Tukey's corrections. * P

    Journal: Nature immunology

    Article Title: A TRAF-like motif of ICOS controls development of germinal center T follicular helper cells via TBK1

    doi: 10.1038/ni.3463

    Figure Lengend Snippet: TBK1 is required for T FH cell differentiation. ( a , c , e ) Flow cytometry of cells from host B6 mice 7 d after adoptive transfer of SMARTA CD4 + T cells transduced with shRNA targeting the Tbk1 (sh Tbk1 -1 and sh Tbk1 -2), Icos or control genes and infected with LCMV Armstrong strain. Numbers adjacent to outlined areas indicate percent CXCR5 + SLAM lo T FH cells ( a ), CXCR5 + PD1 hi GC T FH cells ( c ), or CXCR5 + GL7 hi GC T FH cells ( e ). Cumulative data for a ( b ), c ( d ) or e ( f ) from three independent experiments. ( g ) Flow cytometry of cells from host CD4-Cre × Bcl6 fl/fl mice 10 d after adoptive transfer of Icos –/– SMARTA CD4 + T cells transduced as in ( a ), and were immunized with KLH-gp61 suspended in AddaVax. Numbers adjacent to outlined areas indicate percent CD95 + PNA + GC B cells. Cumulative data for g ( h ) from two independent experiments. ( i ) Quantification of anti-KLH-gp61 IgG from sera of immunized mice analyzed with ELISA and presented as absorbance at 450 nm. The endpoint titer ( j ) and area under curve (AUC; k ) were calculated. Each data point represents a single mouse. Shown are mean ± SEM; ANOVA with post-hoc Tukey's corrections. * P

    Article Snippet: Fluorophore-conjugated anti-CD4 (mouse: clone RM4-5; human: clone RPA-T4), -CD8a (clone 53-6.7), -CD19 (mouse: clone eBio1D3; human: clone HIB19), -CD25 (clone PC61), -CD40L (clone MR1), -CD44 (clone IM7), -CD45.1 (clone A20), -CD62L (clone MEL-14), -GL7 (clone GL-7), -IgD (clone 11-26) and -PD1 (mouse: clone J43; human: clone J105) Abs were obtained from eBioscience, CA.

    Techniques: Cell Differentiation, Flow Cytometry, Cytometry, Mouse Assay, Adoptive Transfer Assay, Transduction, shRNA, Infection, Enzyme-linked Immunosorbent Assay

    Importance of a novel ICOS signaling motif in T FH cell development. ( a ) Evolutionary conservation of the proximal (IProx), PI3K-binding YxxM and distal motifs in the cytoplasmic tail of ICOS. ( b , d ) Flow cytometry of cells from host B6 mice 7 d after adoptive transfer of Icos –/– SMARTA CD4 + T cells transduced with retrovirus encoding empty vector (EV) or wild-type ICOS (WT) or mIProx, YF or TL mutant ICOS and infected with LCMV Armstrong strain. Numbers adjacent to outlined areas indicate percent CXCR5 + SLAM lo T FH cells ( b ) or CXCR5 + PD1 hi GC T FH cells ( d ). Cumulative data for b ( c ) or d ( e ) from three independent experiments. ( f , h ) Flow cytometry of cells from host CD4-Cre × Bcl6 fl/fl mice 10 d after adoptive transfer of Icos –/– SMARTA CD4 + T cells transduced as in ( b ), and were immunized with KLH-gp61 absorbed to alum. Numbers adjacent to outlined areas indicate percent CD95 + GL7 + GC B cells ( f ) and CD138 + IgD – plasma cells ( h ). Cumulative data for f ( g ) or h ( i ) from two independent experiments. Each data point represents a single mouse. ( j ) Quantification of anti-KLH-gp61 IgG from sera of immunized mice analyzed with ELISA and presented as absorbance at 450 nm. The endpoint titer ( k ) and area under curve (AUC; l ) were calculated. Shown are mean ± SEM; ANOVA with post-hoc Tukey's corrections. * P

    Journal: Nature immunology

    Article Title: A TRAF-like motif of ICOS controls development of germinal center T follicular helper cells via TBK1

    doi: 10.1038/ni.3463

    Figure Lengend Snippet: Importance of a novel ICOS signaling motif in T FH cell development. ( a ) Evolutionary conservation of the proximal (IProx), PI3K-binding YxxM and distal motifs in the cytoplasmic tail of ICOS. ( b , d ) Flow cytometry of cells from host B6 mice 7 d after adoptive transfer of Icos –/– SMARTA CD4 + T cells transduced with retrovirus encoding empty vector (EV) or wild-type ICOS (WT) or mIProx, YF or TL mutant ICOS and infected with LCMV Armstrong strain. Numbers adjacent to outlined areas indicate percent CXCR5 + SLAM lo T FH cells ( b ) or CXCR5 + PD1 hi GC T FH cells ( d ). Cumulative data for b ( c ) or d ( e ) from three independent experiments. ( f , h ) Flow cytometry of cells from host CD4-Cre × Bcl6 fl/fl mice 10 d after adoptive transfer of Icos –/– SMARTA CD4 + T cells transduced as in ( b ), and were immunized with KLH-gp61 absorbed to alum. Numbers adjacent to outlined areas indicate percent CD95 + GL7 + GC B cells ( f ) and CD138 + IgD – plasma cells ( h ). Cumulative data for f ( g ) or h ( i ) from two independent experiments. Each data point represents a single mouse. ( j ) Quantification of anti-KLH-gp61 IgG from sera of immunized mice analyzed with ELISA and presented as absorbance at 450 nm. The endpoint titer ( k ) and area under curve (AUC; l ) were calculated. Shown are mean ± SEM; ANOVA with post-hoc Tukey's corrections. * P

    Article Snippet: Fluorophore-conjugated anti-CD4 (mouse: clone RM4-5; human: clone RPA-T4), -CD8a (clone 53-6.7), -CD19 (mouse: clone eBio1D3; human: clone HIB19), -CD25 (clone PC61), -CD40L (clone MR1), -CD44 (clone IM7), -CD45.1 (clone A20), -CD62L (clone MEL-14), -GL7 (clone GL-7), -IgD (clone 11-26) and -PD1 (mouse: clone J43; human: clone J105) Abs were obtained from eBioscience, CA.

    Techniques: Binding Assay, Flow Cytometry, Cytometry, Mouse Assay, Adoptive Transfer Assay, Transduction, Plasmid Preparation, Mutagenesis, Infection, Enzyme-linked Immunosorbent Assay

    Activation of PI3K/Akt signalling regulates neuronal FoxA1 and PDL1. ( a ) Real-time PCR analysed fold changes of Pi3k and Akt mRNA in CGN. Graphs are mean±s.e.m. from three independent experiments. Two-way analysis of variance (ANOVA) test was used, * P

    Journal: Nature Communications

    Article Title: Neuronal IFN-beta–induced PI3K/Akt-FoxA1 signalling is essential for generation of FoxA1+Treg cells

    doi: 10.1038/ncomms14709

    Figure Lengend Snippet: Activation of PI3K/Akt signalling regulates neuronal FoxA1 and PDL1. ( a ) Real-time PCR analysed fold changes of Pi3k and Akt mRNA in CGN. Graphs are mean±s.e.m. from three independent experiments. Two-way analysis of variance (ANOVA) test was used, * P

    Article Snippet: For some experiments, PI3K inhibitor Wortmannin (catalogue number S2758, Selleckchem), Akt inhibitor MK-2206 (catalogue number S1078, Selleckchem), rIFNβ (12405-1, PBL Assay Science) and anti-PDL1 (10 μg ml−1 , Anti-Mouse CD274/B7-H1) Functional Grade Purified, 16-5982-82, eBioscience) were added to the CGNs.

    Techniques: Activation Assay, Real-time Polymerase Chain Reaction

    Generation of FoxA1 + T reg cells requires neuronal FoxA1 and PDL1. ( a ) Percentage of FoxA1 + T regs upon co-culture of T encs with CGNs, with or without transwell. Graphs are mean±s.e.m., n =3. One-way analysis of variance (ANOVA) test was used, *** P

    Journal: Nature Communications

    Article Title: Neuronal IFN-beta–induced PI3K/Akt-FoxA1 signalling is essential for generation of FoxA1+Treg cells

    doi: 10.1038/ncomms14709

    Figure Lengend Snippet: Generation of FoxA1 + T reg cells requires neuronal FoxA1 and PDL1. ( a ) Percentage of FoxA1 + T regs upon co-culture of T encs with CGNs, with or without transwell. Graphs are mean±s.e.m., n =3. One-way analysis of variance (ANOVA) test was used, *** P

    Article Snippet: For some experiments, PI3K inhibitor Wortmannin (catalogue number S2758, Selleckchem), Akt inhibitor MK-2206 (catalogue number S1078, Selleckchem), rIFNβ (12405-1, PBL Assay Science) and anti-PDL1 (10 μg ml−1 , Anti-Mouse CD274/B7-H1) Functional Grade Purified, 16-5982-82, eBioscience) were added to the CGNs.

    Techniques: Co-Culture Assay

    Neuronal ability to generate FoxA1 + T regs is IFNβ-mediated PI3K-Akt-FoxA1 and PDL1 dependent. ( a ) Representative FACS plots of T enc cells after co-culture with CGNs, with and without Akt siRNA KD or PI3K inhibitor (Wortmannin in DMSO, 200 nm). FoxA1 + T reg cells were gated on CD4 high PDL1 high cells. ( b ) FoxA1 expression was gated in CD4 high PDL1 high cells (R1) and in CD4 low PDL1 low cells (R2) and ( c ) quantified percentage of FoxA1 + T regs . Graphs are mean±s.e.m., n =3, one-way analysis of variance (ANOVA) test was used, *** P

    Journal: Nature Communications

    Article Title: Neuronal IFN-beta–induced PI3K/Akt-FoxA1 signalling is essential for generation of FoxA1+Treg cells

    doi: 10.1038/ncomms14709

    Figure Lengend Snippet: Neuronal ability to generate FoxA1 + T regs is IFNβ-mediated PI3K-Akt-FoxA1 and PDL1 dependent. ( a ) Representative FACS plots of T enc cells after co-culture with CGNs, with and without Akt siRNA KD or PI3K inhibitor (Wortmannin in DMSO, 200 nm). FoxA1 + T reg cells were gated on CD4 high PDL1 high cells. ( b ) FoxA1 expression was gated in CD4 high PDL1 high cells (R1) and in CD4 low PDL1 low cells (R2) and ( c ) quantified percentage of FoxA1 + T regs . Graphs are mean±s.e.m., n =3, one-way analysis of variance (ANOVA) test was used, *** P

    Article Snippet: For some experiments, PI3K inhibitor Wortmannin (catalogue number S2758, Selleckchem), Akt inhibitor MK-2206 (catalogue number S1078, Selleckchem), rIFNβ (12405-1, PBL Assay Science) and anti-PDL1 (10 μg ml−1 , Anti-Mouse CD274/B7-H1) Functional Grade Purified, 16-5982-82, eBioscience) were added to the CGNs.

    Techniques: FACS, Co-Culture Assay, Expressing