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

Article Title: Human cytomegalovirus differentially controls B cell and T cell responses through effects on plasmacytoid dendritic cells.

doi: 10.4049/jimmunol.179.11.7767

Figure Lengend Snippet: FIGURE 2. HCMV activates PDCs. A, HCMV alters PDC morphology. PDCs were incubated overnight with medium or TB40/E at an MOI of 5 and examined by light microscopy and Evans blue staining of cytospin preparations. At 24 h, mock-infected PDCs showed plasmacytoid morphology, whereas TB40E-infected PDCs formed clusters, developed dendrites, and had larger cytoplasm. B and C, HCMV induces partial maturation of PDCs. PDCs were mock-infected (mock inf), stimulated with CpG-A (3 g/ml), or infected with TB40/E or VR1814 (MOI of 5) overnight. The stimuli were removed, cells were washed, and fresh medium was added. After 24 h, expression of CD83, HLA-DR, BDCA-2, CD80, and CD86 was measured by FACS. B, Values are means SEM of seven donors in independent experiments. , p 0.05. Y-axis values indicate MFI. C, Open histograms depict cells stained with isotype-matched control Ab. Representative data from one of seven experiments are shown.

Article Snippet: For cell surface staining, we used Abs against CD1a, CD3, CD4, CD8, CD14, CD19, CD20, CD38, CD80, CD86, HLA-DR, and mouse isotype controls (BD Pharmingen); CD83 (Immunotech); and BDCA-2 and BDCA-4 (Miltenyi Biotec).

Techniques: Incubation, Light Microscopy, Staining, Infection, Expressing, Control

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

Article Title: Human cytomegalovirus differentially controls B cell and T cell responses through effects on plasmacytoid dendritic cells.

doi: 10.4049/jimmunol.179.11.7767

Figure Lengend Snippet: FIGURE 4. Soluble factors secreted by HCMV-infected DCs contribute to B cell activation. A and B, PDCs were mock infected or infected with TB40/E (MOI of 5) for 4 h, washed, and resuspended in complete medium for 18 h. Supernatants were collected and filtered through 0.1-m filters to eliminate viral particles. B cells (2.5 105 cells/well) were cultured in 200 l of supernatant from PDC cultures in 96-well round-bottom plates. B cells were unstimulated or stimulated by B cell-receptor ligation (anti-Ig, 10 g/ml). A, After 3 days, B cells were harvested, stained for the B cell markers CD19, CD20, CD38, and CD86, and analyzed by FACS. Values are means SD. , p 0.05 (n 4 donors in independent experiments). B, For proliferation assay, B cells were labeled with 1 M CFSE, incubated with PDC supernatant for 5 days, harvested, and analyzed by FACS. Data from one representative of three donors in independent experiments are shown. C, MDCs were mock infected or infected with TB40/E (MOI of 5) for 4 h, washed, and resuspended in complete medium for 18 h. Supernatants were collected and filtered. B cells (2.5 105 cells/well) were cultured in 200 l of supernatant from MDC cultures. B cells were unstimulated or stimulated by B cell-receptor ligation (anti-Ig, 10 g/ml). After 3 days, B cells were harvested, stained for the B cell markers CD19, CD20, CD38, and CD86, and analyzed by FACS. Values are means SD. , p 0.05 (n 4 donors in independent experiments).

Article Snippet: For cell surface staining, we used Abs against CD1a, CD3, CD4, CD8, CD14, CD19, CD20, CD38, CD80, CD86, HLA-DR, and mouse isotype controls (BD Pharmingen); CD83 (Immunotech); and BDCA-2 and BDCA-4 (Miltenyi Biotec).

Techniques: Infection, Activation Assay, Cell Culture, Ligation, Staining, Proliferation Assay, Labeling, Incubation

Journal: Cell metabolism

Article Title: Ejection of damaged mitochondria and their removal by macrophages ensure efficient thermogenesis in brown adipose tissue

doi: 10.1016/j.cmet.2022.02.016

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: CD86 Antibody, anti-mouse, FITC, Miltenyi Biotec , Miltenyi Biotec , Cat# 130–123-672; RRID:AB_2889633.

Techniques: Immunofluorescence, Recombinant, Cytometry, Purification, shRNA, Cell Culture, Lysis, XF Assay, Staining, Isolation, Transfection, Live Cell Imaging, Mouse Assay, Software, Expressing

Journal: Journal of viral hepatitis

Article Title: A class C CpG toll-like receptor 9 agonist successfully induces robust interferon-alpha production by plasmacytoid dendritic cells from patients chronically infected with hepatitis C.

doi: 10.1111/j.1365-2893.2008.01011.x

Figure Lengend Snippet: Fig. 4 Immunophenotypic maturation of pDCs is not impaired in cHCV infection. After 20–24 h stimulation with TLR agonists, pDCs were processed for flow cytometry to identify immunophenotypic changes consistent with maturation. FACS histograms from a representative NHD and cHCV subject show (a) the rise in CD86 expression and (b) fall in BDCA-2 expression following stimulation with CpG-C (solid histogram) compared to unstimulated cells (open histogram overlay). The bar graphs show the average fold change in mean fluorescence intensity (MFI) ± SEM of (a) CD86 and (b) BDCA-2 on pDCs from 9 NHD (solid bars) and eight cHCV (open bars) subjects (seven NHD for TLR7 agonists) induced by the four TLR agonists. For each TLR agonist, the degree of (a) upregulation of CD86 and (b) downregulation of BDCA-2 did not differ significantly between NHD–pDC and cHCV–pDCs (n.s.). CpG-C was a significantly stronger stimulus for CD86 upregulation than CpG-A (NHD–pDCs, P < 0.001; cHCV–pDCs, P < 0.01) and for cHCV–pDCs loxoribine was also significantly more potent than CpG-A (P < 0.05). CpG-C caused significantly stronger downregulation of BDCA-2 than CpG-A (NHD–pDCs, P < 0.01; cHVC–pDCs, P < 0.01).

Article Snippet: Cells in PBS containing 0.1% NaN3, 1% BSA and 100 lg ⁄ mL human IgG (Jackson Immunoresearch, West Grove, PA, USA) were blocked for 15 min on ice then labelled with monoclonal antibodies against HLA-DR FITC (clone L243), CD86-FITC (clone FUN-1), CD83-APC (clone HB15e) (all from BD Biosciences) or CD303-APC (BDCA2, clone AC144; Miltenyi Biotec) or with isotype controls [murine IgG1 (clone MOPC 31, APC or FITC conjugated) or IgG2a FITC (clone G155-178) (all from BD Biosciences)].

Techniques: Infection, Cytometry, Expressing

Journal: Molecules (Basel, Switzerland)

Article Title: Inosine Prevents Colorectal Cancer Progression by Inducing M1 Phenotypic Polarization of Macrophages.

doi: 10.3390/molecules30010123

Figure Lengend Snippet: Figure 1. Inosine promoted macrophage polarization toward the M1 phenotype. (A) RAW264.7 cells were treated with inosine (0–40 mM) for 24 h; then, an MTT assay was performed to observe cell viability. RAW264.7 cells were administrated with inosine (1.25, 2.5, and 5 mM) in the absence or presence of LPS+IFN-γ or IL-4. (B,C) The proportions of CD86 and CD206-positive cells were determined by a flow cytometer. (D,E) The expression levels of CD86 and CD206 mRNA were detected by RT-qPCR. (F–H) Levels of CD86 and iNOS were measured by WB. Compared to the control group: * p < 0.05, ** p < 0.01, *** p < 0.001; Compared with the LPS+IFN-γ induced M1 group: # p < 0.05, ## p < 0.01, ### p < 0.001; Compared with the IL-4 induced M2 group: ∆∆∆p < 0.001.

Article Snippet: Primary antibodies against CD86 (1:1000, 26903-1-AP, Proteintech, Wuhan, China), iNOS (1:300, 22226-1-AP, Proteintech, Wuhan, China), β-actin (1:2000, GB15003-100, Servicebio, Wuhan, China) and secondary antibody against HRP Goat Anti-Rabbit IgG (1:5000, AS014, ABclonal, Wuhan, China) were used, respectively.

Techniques: MTT Assay, Flow Cytometry, Expressing, Quantitative RT-PCR, Control

Journal: Molecules (Basel, Switzerland)

Article Title: Inosine Prevents Colorectal Cancer Progression by Inducing M1 Phenotypic Polarization of Macrophages.

doi: 10.3390/molecules30010123

Figure Lengend Snippet: Figure 4. Effects of inosine on immune factors in the CT26 tumor microenvironment. (A) Effect of inosine on Ki-67 expression in tumor tissues (Scale: 100 µm; 400× and 200×). (B) Statistics of Ki-67 protein positive expression in tumor tissues (n = 3). (C) Fluorescence co-localization fluorogram of M1- type macrophage marker F4/80 + CD86 (scale: 100 µm; 200×). (D) F4/80 + CD86 expression statistics in tumor tissues. (E) M2 type macrophage marker F4/80 + CD206 fluorescence co-localization fluorogram (scale: 100 µm; 200×). (F) F4/80 + CD206 expression statistics in tumor tissues. Note: 5-Fu are 5-Fu (12 mg/kg) groups. IS-L and IS-H are inosine low and high-dose (5 mg/kg and 50 mg/kg) groups, respectively. Yellow arrows represent positive positions. Compared with the model group: ## p < 0.01; Compared with the 5-Fu group: △p < 0.05.

Article Snippet: Primary antibodies against CD86 (1:1000, 26903-1-AP, Proteintech, Wuhan, China), iNOS (1:300, 22226-1-AP, Proteintech, Wuhan, China), β-actin (1:2000, GB15003-100, Servicebio, Wuhan, China) and secondary antibody against HRP Goat Anti-Rabbit IgG (1:5000, AS014, ABclonal, Wuhan, China) were used, respectively.

Techniques: Expressing, Fluorescence, Marker

Journal: bioRxiv

Article Title: Adjuvant Discovery via a High Throughput Screen using Human Primary Mononuclear Cells

doi: 10.1101/2022.06.17.496630

Figure Lengend Snippet: Key resource table

Article Snippet: CD86 Antibody, anti-human, PE, REAfinityTM (Clone- REA968) , Miltenyi Biotec , Order Number: 130-116-160.

Techniques: Recombinant, Sterility, Injection, Modification, Staining, Saline, Enzyme-linked Immunosorbent Assay, Multiplex Assay, Software

Journal: Pharmaceutics

Article Title: Development and Evaluation of a Novel Diammonium Glycyrrhizinate Phytosome for Nasal Vaccination

doi: 10.3390/pharmaceutics14102000

Figure Lengend Snippet: Flow cytometric analysis of CD11c, CD80, and CD86 expression in BMDCs after treatment of blank medium, free DG, and DG-P for 24 h ( n = 4).

Article Snippet: Fluorochrome-labelled anti-mouse monoclonal antibodies including APC-CD11c, FITC-CD80, and PE-CD86, were purchased from Proteintech (Rosemont, IL, USA).

Techniques: Expressing

Journal: Pharmaceutics

Article Title: Development and Evaluation of a Novel Diammonium Glycyrrhizinate Phytosome for Nasal Vaccination

doi: 10.3390/pharmaceutics14102000

Figure Lengend Snippet: Quantification of CD80 and CD86 expression on BMDCs after activation ( n = 4). Note: *, ** and *** represent the differences at p < 0.05, p < 0.01, and p < 0.001, respectively.

Article Snippet: Fluorochrome-labelled anti-mouse monoclonal antibodies including APC-CD11c, FITC-CD80, and PE-CD86, were purchased from Proteintech (Rosemont, IL, USA).

Techniques: Expressing, Activation Assay

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) Representative flow-cytometry histograms of CD28-huFc staining of the indicated types of Raji cells. Bound CD28-huFc was labeled by AF647 anti-human IgG Fc, the MFI of which was plotted against (CD28-huFc. Shown in gray are Raji (CD80+CD86−) cells stained by isolated huFc domain. Means ± SEM, n = 3.

Article Snippet: The SLBs were then overlaid with 200 μL of either 3 nM human PD-1-His (Sino Biological, 10377-H08H), 3 nM human CD80–His (Sino Biological, 10698-H08H), or 3 nM human CD86–His (Sino Biological, 10699-H08H).

Techniques: Flow Cytometry, Staining, Labeling, Isolation

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) Representative TIRF images of PD-L1 LUVs captured by PD-1 SLB, CD80 SLB, or CD86 SLB; each LUV is registered as a green spot. Bar graph summarizes the fluorescence intensity (FI) of the LUV channel under indicated conditions, normalized to the intensity of the condition with PD-1 SLBs. Data are means ± SEM, n = 3. Scale bars, 5 μm.

Article Snippet: The SLBs were then overlaid with 200 μL of either 3 nM human PD-1-His (Sino Biological, 10377-H08H), 3 nM human CD80–His (Sino Biological, 10698-H08H), or 3 nM human CD86–His (Sino Biological, 10699-H08H).

Techniques: Fluorescence

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) Representative flow-cytometry histograms of CTLA-4-huFc staining of the indicated types of Raji cells. Bound CTLA-4-huFc was labeled by AF647 anti-human IgG Fc, the MFI of which was plotted against (CTLA-4-huFc). Shown in gray are Raji (CD80+CD86−) cells stained by isolated huFc domain. Means ± SEM, n ≥ 3.

Article Snippet: The SLBs were then overlaid with 200 μL of either 3 nM human PD-1-His (Sino Biological, 10377-H08H), 3 nM human CD80–His (Sino Biological, 10698-H08H), or 3 nM human CD86–His (Sino Biological, 10699-H08H).

Techniques: Flow Cytometry, Staining, Labeling, Isolation

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) On the left are flow-cytometry histograms of CD80 and CD86 surface levels on DCs and macrophages (Macs) isolated from tumor tissues of 4T1 implanted BALB/C female mice treated with anti-PD-L1 (magenta traces), anti-PD-1 (cyan traces), or control IgG (black traces). On the right are bar graphs summarizing the MFI of CD80 and CD86 staining under the indicated conditions. Data are shown as mean ± SEM, n ≥ 3 mice.

Article Snippet: The SLBs were then overlaid with 200 μL of either 3 nM human PD-1-His (Sino Biological, 10377-H08H), 3 nM human CD80–His (Sino Biological, 10698-H08H), or 3 nM human CD86–His (Sino Biological, 10699-H08H).

Techniques: Flow Cytometry, Isolation, Staining

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: The SLBs were then overlaid with 200 μL of either 3 nM human PD-1-His (Sino Biological, 10377-H08H), 3 nM human CD80–His (Sino Biological, 10698-H08H), or 3 nM human CD86–His (Sino Biological, 10699-H08H).

Techniques: Recombinant, Isolation, Enzyme-linked Immunosorbent Assay, Cell Isolation, Software, Imaging