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cd160 antibody 688327  (Jackson Immuno)
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Jackson Immuno cd160 antibody 688327
Regulation of HVEM co-stimulation in trans and in cis. (A) Upper panels: cell surface expression of CD86, <t>CD160</t> and LIGHT on TCS analysed via flow cytometry (open histograms: control TCS; grey histograms: TCS expressing the indicated molecules). Lower panel: membrane-bound anti-CD3-fragment (detected via its CD14 stem) on TCS analysed via flow cytometry. Parental BW5147 cells were used as a control. (B) Control reporter cells and HVEM expressing reporter cells were stimulated with control TCS and TCS expressing CD86, BTLA, LIGHT and CD160 for 24 h and NFκB::eGFP expression was measured by flow cytometry. Data is shown from 4 independent experiments performed in duplicates. For statistical evaluation, one-way ANOVA with Dunn’s multiple-comparisons test was performed (***p ≤ 0.001; **p ≤ 0.01; *p < 0.05; ns, p > 0.05). (C) Jurkat NFκB::eGFP reporter cells expressing HVEM, HVEM/BTLA, HVEM/LIGHT and HVEM/CD160 were analysed for the expression of the indicated molecules; open histogram show control reporter cells. (D) The indicated reporter cells were left unstimulated or were stimulated with control TCS or TCS CD86 and eGFP expression was measured via flow cytometry. Results are shown from 4 independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; ns, p > 0.05). (E) Expression levels of HVEM and BTLAΔcyt on reporter cells. Open histograms represent control reporter cells. (F) The indicated reporter cells were left unstimulated or were co-cultured with control TCS and TCS CD86. eGFP expression was assessed via flow cytometry. Data is shown for 4 independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; **p ≤ 0.01; ns, p > 0.05). (B, D, F) ± SD is shown.
Cd160 Antibody 688327, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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cd160 antibody 688327 - by Bioz Stars, 2026-09
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Regulation of HVEM co-stimulation in trans and in cis. (A) Upper panels: cell surface expression of CD86, CD160 and LIGHT on TCS analysed via flow cytometry (open histograms: control TCS; grey histograms: TCS expressing the indicated molecules). Lower panel: membrane-bound anti-CD3-fragment (detected via its CD14 stem) on TCS analysed via flow cytometry. Parental BW5147 cells were used as a control. (B) Control reporter cells and HVEM expressing reporter cells were stimulated with control TCS and TCS expressing CD86, BTLA, LIGHT and CD160 for 24 h and NFκB::eGFP expression was measured by flow cytometry. Data is shown from 4 independent experiments performed in duplicates. For statistical evaluation, one-way ANOVA with Dunn’s multiple-comparisons test was performed (***p ≤ 0.001; **p ≤ 0.01; *p < 0.05; ns, p > 0.05). (C) Jurkat NFκB::eGFP reporter cells expressing HVEM, HVEM/BTLA, HVEM/LIGHT and HVEM/CD160 were analysed for the expression of the indicated molecules; open histogram show control reporter cells. (D) The indicated reporter cells were left unstimulated or were stimulated with control TCS or TCS CD86 and eGFP expression was measured via flow cytometry. Results are shown from 4 independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; ns, p > 0.05). (E) Expression levels of HVEM and BTLAΔcyt on reporter cells. Open histograms represent control reporter cells. (F) The indicated reporter cells were left unstimulated or were co-cultured with control TCS and TCS CD86. eGFP expression was assessed via flow cytometry. Data is shown for 4 independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; **p ≤ 0.01; ns, p > 0.05). (B, D, F) ± SD is shown.

Journal: Frontiers in Immunology

Article Title: BTLA inhibition has a dominant role in the cis -complex of BTLA and HVEM

doi: 10.3389/fimmu.2022.956694

Figure Lengend Snippet: Regulation of HVEM co-stimulation in trans and in cis. (A) Upper panels: cell surface expression of CD86, CD160 and LIGHT on TCS analysed via flow cytometry (open histograms: control TCS; grey histograms: TCS expressing the indicated molecules). Lower panel: membrane-bound anti-CD3-fragment (detected via its CD14 stem) on TCS analysed via flow cytometry. Parental BW5147 cells were used as a control. (B) Control reporter cells and HVEM expressing reporter cells were stimulated with control TCS and TCS expressing CD86, BTLA, LIGHT and CD160 for 24 h and NFκB::eGFP expression was measured by flow cytometry. Data is shown from 4 independent experiments performed in duplicates. For statistical evaluation, one-way ANOVA with Dunn’s multiple-comparisons test was performed (***p ≤ 0.001; **p ≤ 0.01; *p < 0.05; ns, p > 0.05). (C) Jurkat NFκB::eGFP reporter cells expressing HVEM, HVEM/BTLA, HVEM/LIGHT and HVEM/CD160 were analysed for the expression of the indicated molecules; open histogram show control reporter cells. (D) The indicated reporter cells were left unstimulated or were stimulated with control TCS or TCS CD86 and eGFP expression was measured via flow cytometry. Results are shown from 4 independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; ns, p > 0.05). (E) Expression levels of HVEM and BTLAΔcyt on reporter cells. Open histograms represent control reporter cells. (F) The indicated reporter cells were left unstimulated or were co-cultured with control TCS and TCS CD86. eGFP expression was assessed via flow cytometry. Data is shown for 4 independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; **p ≤ 0.01; ns, p > 0.05). (B, D, F) ± SD is shown.

Article Snippet: Binding of the CD160 antibody 688327 was detected with PE- labelled goat-anti mouse-IgG (Fc specific) antibodies (Jackson ImmunoResearch, West Grove, PA, USA).

Techniques: Expressing, Flow Cytometry, Cell Culture

Assessment of co-expression of HVEM/BTLA signaling. (A) Control reporter cells and reporter cells expressing BTLA, HVEM or BTLA/HVEM were left unstimulated or stimulated with TCS ctrl, TCS BTLA and TCS HVEM. NF-κB::eGFP activation was measured after 24 h. Left: one representative experiment performed in duplicate is shown. Right: reporter gene expression is shown normalized to reporter gene expression induced by control-TCS (gMFI of reporter cells stimulated with the indicated TCS/gMFI of TCS ctrl stimulated cells). Data is depicted of at least eight independent experiments performed in duplicates. One-way analysis of variance followed by a Dunnett’s multiple comparison test were used for comparison to control reporter cells (***p ≤ 0.001; ns, p > 0.05). (B) Jurkat NFκB::eGFP ctrl and Jurkat NFκB::eGFP expressing HVEM, HVEM/BTLA or HVEM/BTLAΔcyt cells were stimulated with TCS ctrl, TCS CD86, TCS CD160 and TCS LIGHT. Reporter gene expression is shown normalized to control-TCS. Results are shown from three independent experiments performed in duplicates. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001). (C) Left panel: Schematic representation of receptors on reporter cells and ligands on stimulator that were evaluated in absence or presence of a blocking BTLA antibody. Right panel: Reporter cells expressing BTLA/HVEM were stimulated with TCS CD86, TCS CD160 or TCS LIGHT in the presence or absence of a blocking BTLA antibody (5 μg/ml; clone 6F4). Reporter activation induced by the indicated TCS is shown normalized to control-TCS. Data is depicted from three independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; ns, p > 0.05). (D) Flow cytometric analysis of cell surface molecules BTLA and HVEM on the indicated reporter cells. (E) Left panel: Reporter cells expressing HVEM, HVEM/BTLA high or HVEM/BTLA low were left unstimulated or stimulated with TCS ctrl, TCS BTLA and TCS LIGHT. Right panel: Reporter cells expressing HVEM, HVEM/BTLA high or HVEM/BTLA low were left unstimulated or stimulated with TCS ctrl or TCS HVEM. gMFI of NFκB::eGFP activation is shown. Results are depicted from four independent experiments performed in duplicate. (F) Cell surface expression of mHVEM and membrane-bound anti-CD3 on TCS analysed via flow cytometry (open histograms: control TCS; grey histograms: expression level of the indicated molecules). (G) Left panel: Schematic representation of stimulation experiments of reporter cells co-expressing BTLA and HVEM with TCS expressing human HVEM or mouse HVEM (mHVEM) with and without HVEM antibody SL030717 (10 µg/ml) that blocks human but not mouse HVEM. Right panel: Control reporter cells and reporter cells expressing HVEM/BTLA or BTLA were stimulated with TCS HVEM or TCS mHVEM in the presence or absence of a blocking HVEM antibody. Reporter gene expression is shown normalized to control-TCS. Results are depicted from four independent experiments performed in duplicate. (A, B, C, G) ±ΔSD is shown. Dotted line depicts control stimulation.

Journal: Frontiers in Immunology

Article Title: BTLA inhibition has a dominant role in the cis -complex of BTLA and HVEM

doi: 10.3389/fimmu.2022.956694

Figure Lengend Snippet: Assessment of co-expression of HVEM/BTLA signaling. (A) Control reporter cells and reporter cells expressing BTLA, HVEM or BTLA/HVEM were left unstimulated or stimulated with TCS ctrl, TCS BTLA and TCS HVEM. NF-κB::eGFP activation was measured after 24 h. Left: one representative experiment performed in duplicate is shown. Right: reporter gene expression is shown normalized to reporter gene expression induced by control-TCS (gMFI of reporter cells stimulated with the indicated TCS/gMFI of TCS ctrl stimulated cells). Data is depicted of at least eight independent experiments performed in duplicates. One-way analysis of variance followed by a Dunnett’s multiple comparison test were used for comparison to control reporter cells (***p ≤ 0.001; ns, p > 0.05). (B) Jurkat NFκB::eGFP ctrl and Jurkat NFκB::eGFP expressing HVEM, HVEM/BTLA or HVEM/BTLAΔcyt cells were stimulated with TCS ctrl, TCS CD86, TCS CD160 and TCS LIGHT. Reporter gene expression is shown normalized to control-TCS. Results are shown from three independent experiments performed in duplicates. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001). (C) Left panel: Schematic representation of receptors on reporter cells and ligands on stimulator that were evaluated in absence or presence of a blocking BTLA antibody. Right panel: Reporter cells expressing BTLA/HVEM were stimulated with TCS CD86, TCS CD160 or TCS LIGHT in the presence or absence of a blocking BTLA antibody (5 μg/ml; clone 6F4). Reporter activation induced by the indicated TCS is shown normalized to control-TCS. Data is depicted from three independent experiments performed in duplicate. For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; ns, p > 0.05). (D) Flow cytometric analysis of cell surface molecules BTLA and HVEM on the indicated reporter cells. (E) Left panel: Reporter cells expressing HVEM, HVEM/BTLA high or HVEM/BTLA low were left unstimulated or stimulated with TCS ctrl, TCS BTLA and TCS LIGHT. Right panel: Reporter cells expressing HVEM, HVEM/BTLA high or HVEM/BTLA low were left unstimulated or stimulated with TCS ctrl or TCS HVEM. gMFI of NFκB::eGFP activation is shown. Results are depicted from four independent experiments performed in duplicate. (F) Cell surface expression of mHVEM and membrane-bound anti-CD3 on TCS analysed via flow cytometry (open histograms: control TCS; grey histograms: expression level of the indicated molecules). (G) Left panel: Schematic representation of stimulation experiments of reporter cells co-expressing BTLA and HVEM with TCS expressing human HVEM or mouse HVEM (mHVEM) with and without HVEM antibody SL030717 (10 µg/ml) that blocks human but not mouse HVEM. Right panel: Control reporter cells and reporter cells expressing HVEM/BTLA or BTLA were stimulated with TCS HVEM or TCS mHVEM in the presence or absence of a blocking HVEM antibody. Reporter gene expression is shown normalized to control-TCS. Results are depicted from four independent experiments performed in duplicate. (A, B, C, G) ±ΔSD is shown. Dotted line depicts control stimulation.

Article Snippet: Binding of the CD160 antibody 688327 was detected with PE- labelled goat-anti mouse-IgG (Fc specific) antibodies (Jackson ImmunoResearch, West Grove, PA, USA).

Techniques: Expressing, Activation Assay, Blocking Assay, Flow Cytometry

Evaluation of CD160 receptor function in a triple parameter reporter cell system. (A) Left panel: HVEM expressing triple parameter reporter cells (TPR; grey histogram) and control-TPR (open histogram) were analysed for HVEM expression. Right panel: Control TPR and TPR expressing HVEM were stimulated with TCS control or TCS expressing BTLA, CD160 or LIGHT. Reporter activation (NFAT::eGFP, NFkB::eCFP and AP-1::mCherry) was assessed via flow cytometry. Data of three independent experiments in duplicate is shown. Normalized reporter activation is shown (gMFI reporter gene expression induced by the indicated TCS/gMFI reporter gene expression induced by TCS ctrl stimulated cells). (B) Left panel: BTLA expressing triple parameter reporter cells (TPR; grey histogram) and control-TPR (open histogram) were analysed for BTLA expression. Right panel: Control TPR and TPR expressing BTLA were stimulated with control TCS and TCS HVEM. Results are shown from three independent performed experiments in duplicate. Normalized reporter activation is shown (gMFI of TCS HVEM of stimulated cells/gMFI of TCS ctrl stimulated cells). (A, B) For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; **p ≤ 0.01; *p < 0.05). (C) Left panel: Schematic representation of two CD160 isoforms: the glycosylphosphatidylinositol-anchored-CD160 (CD160-GPI) and the transmembrane isoform of CD160 (CD160-TM). Right panel: Cell surface expression of CD160-GPI and CD160-TM on Jurkat TPR cells assessed by using two different antibodies (clone BY55 and clone 688327) (open histogram: control cells; grey histograms: expression of the indicated CD160 molecules). (D) Control TPR and TPR expressing the two isoforms CD160-GPI and CD160-TM, respectively, were left unstimulated or stimulated with the indicated TCS. (E) Left panel: Schematic representation of a mICOS-CD160-TM chimera. Middle panel: Expression of mICOS chimera on TPR. Right panel: TCS mICOSL (grey histogram) and control TCS (open histogram) were stained with a mICOSL antibody (F) mICOS-chimera expressing reporter cells were stimulated with TCS control and TCS mICOSL. Reporter gene expression induced by TCS mICOSL normalized to TCS control. For statistical evaluation, one-way ANOVA with Dunn’s multiple-comparisons test was performed (***p ≤ 0.001; **p ≤ 0.01; *p < 0.05). (A, B, D, F) ±ΔSD or mean is shown. Dotted line depicts control stimulation.

Journal: Frontiers in Immunology

Article Title: BTLA inhibition has a dominant role in the cis -complex of BTLA and HVEM

doi: 10.3389/fimmu.2022.956694

Figure Lengend Snippet: Evaluation of CD160 receptor function in a triple parameter reporter cell system. (A) Left panel: HVEM expressing triple parameter reporter cells (TPR; grey histogram) and control-TPR (open histogram) were analysed for HVEM expression. Right panel: Control TPR and TPR expressing HVEM were stimulated with TCS control or TCS expressing BTLA, CD160 or LIGHT. Reporter activation (NFAT::eGFP, NFkB::eCFP and AP-1::mCherry) was assessed via flow cytometry. Data of three independent experiments in duplicate is shown. Normalized reporter activation is shown (gMFI reporter gene expression induced by the indicated TCS/gMFI reporter gene expression induced by TCS ctrl stimulated cells). (B) Left panel: BTLA expressing triple parameter reporter cells (TPR; grey histogram) and control-TPR (open histogram) were analysed for BTLA expression. Right panel: Control TPR and TPR expressing BTLA were stimulated with control TCS and TCS HVEM. Results are shown from three independent performed experiments in duplicate. Normalized reporter activation is shown (gMFI of TCS HVEM of stimulated cells/gMFI of TCS ctrl stimulated cells). (A, B) For statistical evaluation, two-way ANOVA followed by Bonferroni’s test was performed (***p ≤ 0.001; **p ≤ 0.01; *p < 0.05). (C) Left panel: Schematic representation of two CD160 isoforms: the glycosylphosphatidylinositol-anchored-CD160 (CD160-GPI) and the transmembrane isoform of CD160 (CD160-TM). Right panel: Cell surface expression of CD160-GPI and CD160-TM on Jurkat TPR cells assessed by using two different antibodies (clone BY55 and clone 688327) (open histogram: control cells; grey histograms: expression of the indicated CD160 molecules). (D) Control TPR and TPR expressing the two isoforms CD160-GPI and CD160-TM, respectively, were left unstimulated or stimulated with the indicated TCS. (E) Left panel: Schematic representation of a mICOS-CD160-TM chimera. Middle panel: Expression of mICOS chimera on TPR. Right panel: TCS mICOSL (grey histogram) and control TCS (open histogram) were stained with a mICOSL antibody (F) mICOS-chimera expressing reporter cells were stimulated with TCS control and TCS mICOSL. Reporter gene expression induced by TCS mICOSL normalized to TCS control. For statistical evaluation, one-way ANOVA with Dunn’s multiple-comparisons test was performed (***p ≤ 0.001; **p ≤ 0.01; *p < 0.05). (A, B, D, F) ±ΔSD or mean is shown. Dotted line depicts control stimulation.

Article Snippet: Binding of the CD160 antibody 688327 was detected with PE- labelled goat-anti mouse-IgG (Fc specific) antibodies (Jackson ImmunoResearch, West Grove, PA, USA).

Techniques: Expressing, Activation Assay, Flow Cytometry, Staining

HVEM engagement in primary human T cells. (A) Representative contour plots of CFSE-labelled PBMCs stimulated with TCS ctrl, TCS CD86, TCS 4-1BBL, TCS BTLA, TCS HVEM, TCS CD160 and TCS LIGHT for 5 days. Percentages of proliferated (CFSE low ) of CD4 + and CD8 + T cells is shown. (B, C) CFSE-labelled PBMCs from healthy donors were stimulated for 5 days with the indicated TCS. Proliferation (CFSE low ) and CD25 upregulation of CD4 + and CD8 + T cells was measured at day 5. Data is normalized for each donor to stimulation with TCS ctrl. Each data point represents the mean of triplicate measurement of one donor (n = 15; TCS 4-1BBL n = 7). (D) Cell culture supernatants of PBMCs stimulated with TCS cells were harvested at day 5 and cytokine expression profile (IFN-γ, GM-CSF and TNF-α) was measured via Luminex multiplex cytokine analysis. Each data point represents the mean of triplicate measurement of one donor (IFN-γ: n = 8, TCS 4-1BBL n = 5; GM-CSF and TNF-α: n = 9, TCS 4-1BBL n = 6). (B-D) For statistical evaluation, a one-way ANOVA with Dunn’s multiple-comparisons test was performed (***p ≤ 0.001; **p ≤ 0.01). Median is shown (red line). Dotted line depicts control stimulation.

Journal: Frontiers in Immunology

Article Title: BTLA inhibition has a dominant role in the cis -complex of BTLA and HVEM

doi: 10.3389/fimmu.2022.956694

Figure Lengend Snippet: HVEM engagement in primary human T cells. (A) Representative contour plots of CFSE-labelled PBMCs stimulated with TCS ctrl, TCS CD86, TCS 4-1BBL, TCS BTLA, TCS HVEM, TCS CD160 and TCS LIGHT for 5 days. Percentages of proliferated (CFSE low ) of CD4 + and CD8 + T cells is shown. (B, C) CFSE-labelled PBMCs from healthy donors were stimulated for 5 days with the indicated TCS. Proliferation (CFSE low ) and CD25 upregulation of CD4 + and CD8 + T cells was measured at day 5. Data is normalized for each donor to stimulation with TCS ctrl. Each data point represents the mean of triplicate measurement of one donor (n = 15; TCS 4-1BBL n = 7). (D) Cell culture supernatants of PBMCs stimulated with TCS cells were harvested at day 5 and cytokine expression profile (IFN-γ, GM-CSF and TNF-α) was measured via Luminex multiplex cytokine analysis. Each data point represents the mean of triplicate measurement of one donor (IFN-γ: n = 8, TCS 4-1BBL n = 5; GM-CSF and TNF-α: n = 9, TCS 4-1BBL n = 6). (B-D) For statistical evaluation, a one-way ANOVA with Dunn’s multiple-comparisons test was performed (***p ≤ 0.001; **p ≤ 0.01). Median is shown (red line). Dotted line depicts control stimulation.

Article Snippet: Binding of the CD160 antibody 688327 was detected with PE- labelled goat-anti mouse-IgG (Fc specific) antibodies (Jackson ImmunoResearch, West Grove, PA, USA).

Techniques: Cell Culture, Expressing, Luminex, Multiplex Assay