apc anti il 1b  (Miltenyi Biotec)

Journal: Journal of Inflammation Research

Article Title: Role of Chemotaxis of Vδ2 T Cells to the Synovium in the Pathogenesis of Acute Gouty Arthritis

doi: 10.2147/JIR.S449329

Figure Lengend Snippet: Clinical Characteristics of Gout Patients and Healthy Controls

Article Snippet: Fluorescein isothiocyanate (FITC)-conjugated anti-human CD14, phycoerythrin (PE)-conjugated anti-human CD16, allophycocyanine (APC)-conjugated anti-human CD192 (CCR2), PE-conjugated anti-human CD294 (CRTH2), APC-conjugated anti-human CD127 (IL-7Rα), peridinin-chlorophyll-protein/cyanine5.5 (PerCP/Cy5.5) – conjugated anti-human CD117 (c-kit), FITC-conjugated anti-human Lineage Cocktail (CD3, CD14, CD19, CD20, CD56), PerCP/Cy5.5-conjugated anti-human CD3, APC-conjugated anti-human CD57, PerCP-conjugated anti-human CD8, PE/Cy7-conjugated anti-human CD56 (NCAM), FITC-conjugated anti-human CD3, PE-conjugated anti-human TCR γ/δ, APC anti-human TCR Vδ2, PerCP/Cy5.5-conjugated anti-human CD3, FITC-conjugated anti-human TCR Vδ2, APC-conjugated anti-human IL-1β, APC-conjugated anti-human IL-17, APC-conjugated anti-human tumor necrosis factor (TNF) -α, APC-conjugated anti-human interferon (IFN) -γ, PE/Cy7-conjugated anti-human C-X-C chemokine receptor 3 (CXCR3), and APC/Fire 750-conjugated anti-human C-C chemokine receptor 5 (CCR5) antibodies were purchased from BD Biosciences (Franklin Lakes, NJ, USA).

Techniques:

Journal: Journal of Inflammation Research

Article Title: Role of Chemotaxis of Vδ2 T Cells to the Synovium in the Pathogenesis of Acute Gouty Arthritis

doi: 10.2147/JIR.S449329

Figure Lengend Snippet: Patients with AGA had low levels of peripheral Vδ2 T cells. ( A ) Illustrated in this figure are representative scatter plots delineating the gating strategy employed for the detection of γδ T cells, Vδ1 T cells, and Vδ2 T cells. Lymphocytes are identified by their forward scatter (FSC) and side scatter (SSC) characteristics, with single cells isolated based on side scatter area (SSC-A) and side scatter height (SSC-H) parameters. Subsequent gating is conducted for γδ T cells using CD3 and TCR γδ markers, and for Vδ1 and Vδ2 T cells using Vδ1 and Vδ2 markers, respectively. ( B-C ) Peripheral blood mononuclear cells derived from HCs and patients with AGA were subjected to staining with anti-CD3, anti-γδ TCR, anti-Vδ1 or anti-Vδ2 monoclonal antibodies (mAbs) for subsequent flow cytometry analysis. The left panels show representative flow cytometry profiles of γδ T cells ( B ), and Vδ1 T cells and Vδ2 T cells ( C ). The right panels show bar graphs of the percentage of the respective cell populations. The sample size consists of n=18 for the HC group, and n=27 for the AGA group. ( D ) The percentage of peripheral Vδ2 T cells in AGA shows a negative correlation with the concentration of CRP, value of ESR and the count of neutrophils (n=21). ( E-G ) Depicted herein are alternations in the percentages of γδ T cells, Vδ2 T cells and Vδ1 T cells across HCs and different stages of gout. Data from HC (n=18), acute period (n=27), remission period (n=11) and intercritical period (n=26) are shown. Results are expressed as mean±SEM. ns, no significance; * P <0.05, and *** P <0.001 signify significance as determined through two-tailed unpaired t -test. Correlations are calculated using Spearman correlation analysis.

Article Snippet: Fluorescein isothiocyanate (FITC)-conjugated anti-human CD14, phycoerythrin (PE)-conjugated anti-human CD16, allophycocyanine (APC)-conjugated anti-human CD192 (CCR2), PE-conjugated anti-human CD294 (CRTH2), APC-conjugated anti-human CD127 (IL-7Rα), peridinin-chlorophyll-protein/cyanine5.5 (PerCP/Cy5.5) – conjugated anti-human CD117 (c-kit), FITC-conjugated anti-human Lineage Cocktail (CD3, CD14, CD19, CD20, CD56), PerCP/Cy5.5-conjugated anti-human CD3, APC-conjugated anti-human CD57, PerCP-conjugated anti-human CD8, PE/Cy7-conjugated anti-human CD56 (NCAM), FITC-conjugated anti-human CD3, PE-conjugated anti-human TCR γ/δ, APC anti-human TCR Vδ2, PerCP/Cy5.5-conjugated anti-human CD3, FITC-conjugated anti-human TCR Vδ2, APC-conjugated anti-human IL-1β, APC-conjugated anti-human IL-17, APC-conjugated anti-human tumor necrosis factor (TNF) -α, APC-conjugated anti-human interferon (IFN) -γ, PE/Cy7-conjugated anti-human C-X-C chemokine receptor 3 (CXCR3), and APC/Fire 750-conjugated anti-human C-C chemokine receptor 5 (CCR5) antibodies were purchased from BD Biosciences (Franklin Lakes, NJ, USA).

Techniques: Isolation, Derivative Assay, Staining, Flow Cytometry, Concentration Assay, Two Tailed Test

Journal: Journal of Inflammation Research

Article Title: Role of Chemotaxis of Vδ2 T Cells to the Synovium in the Pathogenesis of Acute Gouty Arthritis

doi: 10.2147/JIR.S449329

Figure Lengend Snippet: The Proportions of γδT Cells and Their Subtypes in Gout Patients and Healthy Controls

Article Snippet: Fluorescein isothiocyanate (FITC)-conjugated anti-human CD14, phycoerythrin (PE)-conjugated anti-human CD16, allophycocyanine (APC)-conjugated anti-human CD192 (CCR2), PE-conjugated anti-human CD294 (CRTH2), APC-conjugated anti-human CD127 (IL-7Rα), peridinin-chlorophyll-protein/cyanine5.5 (PerCP/Cy5.5) – conjugated anti-human CD117 (c-kit), FITC-conjugated anti-human Lineage Cocktail (CD3, CD14, CD19, CD20, CD56), PerCP/Cy5.5-conjugated anti-human CD3, APC-conjugated anti-human CD57, PerCP-conjugated anti-human CD8, PE/Cy7-conjugated anti-human CD56 (NCAM), FITC-conjugated anti-human CD3, PE-conjugated anti-human TCR γ/δ, APC anti-human TCR Vδ2, PerCP/Cy5.5-conjugated anti-human CD3, FITC-conjugated anti-human TCR Vδ2, APC-conjugated anti-human IL-1β, APC-conjugated anti-human IL-17, APC-conjugated anti-human tumor necrosis factor (TNF) -α, APC-conjugated anti-human interferon (IFN) -γ, PE/Cy7-conjugated anti-human C-X-C chemokine receptor 3 (CXCR3), and APC/Fire 750-conjugated anti-human C-C chemokine receptor 5 (CCR5) antibodies were purchased from BD Biosciences (Franklin Lakes, NJ, USA).

Techniques:

Journal: Journal of Inflammation Research

Article Title: Role of Chemotaxis of Vδ2 T Cells to the Synovium in the Pathogenesis of Acute Gouty Arthritis

doi: 10.2147/JIR.S449329

Figure Lengend Snippet: Vδ2 T cells accumulated in gout synovial fluid. ( A and B ) The proportions of γδ T cells within the lymphocytes subset ( A ) and the ratios of Vδ2 T cells and Vδ1 T cells among the total γδ T cells populations ( B ) were quantified through flow cytometry analyses for OA-SFMC samples (n=9) and AGA-SFMC samples (n=9). Representative flow profiles are shown and the percentages were summarized. ( C and D ) The left panels show representative flow profiles of γδ T cells ( C ), Vδ1 T cells and Vδ2 T cells ( D ) derived from both PBMC, and SFMC from patients with gout (n=8). The right panels show bar graphs of the percentage of corresponding cells within the indicated populations. Results are expressed as mean±SEM. ns, no significance; ** P <0.01, and *** P <0.001 signify significance as determined through two-tailed paired t -test.

Article Snippet: Fluorescein isothiocyanate (FITC)-conjugated anti-human CD14, phycoerythrin (PE)-conjugated anti-human CD16, allophycocyanine (APC)-conjugated anti-human CD192 (CCR2), PE-conjugated anti-human CD294 (CRTH2), APC-conjugated anti-human CD127 (IL-7Rα), peridinin-chlorophyll-protein/cyanine5.5 (PerCP/Cy5.5) – conjugated anti-human CD117 (c-kit), FITC-conjugated anti-human Lineage Cocktail (CD3, CD14, CD19, CD20, CD56), PerCP/Cy5.5-conjugated anti-human CD3, APC-conjugated anti-human CD57, PerCP-conjugated anti-human CD8, PE/Cy7-conjugated anti-human CD56 (NCAM), FITC-conjugated anti-human CD3, PE-conjugated anti-human TCR γ/δ, APC anti-human TCR Vδ2, PerCP/Cy5.5-conjugated anti-human CD3, FITC-conjugated anti-human TCR Vδ2, APC-conjugated anti-human IL-1β, APC-conjugated anti-human IL-17, APC-conjugated anti-human tumor necrosis factor (TNF) -α, APC-conjugated anti-human interferon (IFN) -γ, PE/Cy7-conjugated anti-human C-X-C chemokine receptor 3 (CXCR3), and APC/Fire 750-conjugated anti-human C-C chemokine receptor 5 (CCR5) antibodies were purchased from BD Biosciences (Franklin Lakes, NJ, USA).

Techniques: Flow Cytometry, Derivative Assay, Two Tailed Test

Journal: Journal of Inflammation Research

Article Title: Role of Chemotaxis of Vδ2 T Cells to the Synovium in the Pathogenesis of Acute Gouty Arthritis

doi: 10.2147/JIR.S449329

Figure Lengend Snippet: CXCR3 and CXCL10 may be associated with the chemotaxis of Vδ2 T cell in AGA. ( A ) Transwell migration assay demonstrated a pronounced capacity of chemotaxis in Vδ2 T cells isolated from AGA patients. Data were pooled from three independent experiments. ( B ) Comparison of the proportion of CXCR3-positive cells in Vδ2 T cells from HCs and AGA patients. ( C ) Comparison of the proportion of CXCR3-positive cells in Vδ2 T cells and Vδ1 T cells from AGA patients. ( D ) Comparison of the proportion of CCR5-positive cells in Vδ2 T cells from HCs and AGA patients. Data of HC (n=8) and AGA (n=10) are shown. ( E and F ) The concentrations of established ligands of CXCR3 and CCR5 were quantified within SF of gout patients (n=18), RA patients (n=7), OA patients (n=7), and serum of gout patients (n=49). Results are expressed as mean±SEM. * P <0.05, ** P <0.01, and *** P <0.001 signify significance as determined through the Mann–Whitney U -test ( A and E ) and two-tailed unpaired t -test ( B–D ).

Article Snippet: Fluorescein isothiocyanate (FITC)-conjugated anti-human CD14, phycoerythrin (PE)-conjugated anti-human CD16, allophycocyanine (APC)-conjugated anti-human CD192 (CCR2), PE-conjugated anti-human CD294 (CRTH2), APC-conjugated anti-human CD127 (IL-7Rα), peridinin-chlorophyll-protein/cyanine5.5 (PerCP/Cy5.5) – conjugated anti-human CD117 (c-kit), FITC-conjugated anti-human Lineage Cocktail (CD3, CD14, CD19, CD20, CD56), PerCP/Cy5.5-conjugated anti-human CD3, APC-conjugated anti-human CD57, PerCP-conjugated anti-human CD8, PE/Cy7-conjugated anti-human CD56 (NCAM), FITC-conjugated anti-human CD3, PE-conjugated anti-human TCR γ/δ, APC anti-human TCR Vδ2, PerCP/Cy5.5-conjugated anti-human CD3, FITC-conjugated anti-human TCR Vδ2, APC-conjugated anti-human IL-1β, APC-conjugated anti-human IL-17, APC-conjugated anti-human tumor necrosis factor (TNF) -α, APC-conjugated anti-human interferon (IFN) -γ, PE/Cy7-conjugated anti-human C-X-C chemokine receptor 3 (CXCR3), and APC/Fire 750-conjugated anti-human C-C chemokine receptor 5 (CCR5) antibodies were purchased from BD Biosciences (Franklin Lakes, NJ, USA).

Techniques: Chemotaxis Assay, Transwell Migration Assay, Isolation, Comparison, MANN-WHITNEY, Two Tailed Test

Journal: Journal of Inflammation Research

Article Title: Role of Chemotaxis of Vδ2 T Cells to the Synovium in the Pathogenesis of Acute Gouty Arthritis

doi: 10.2147/JIR.S449329

Figure Lengend Snippet: Vδ2 T cells in patients with AGA secreted high levels of IL-17. ( A ) Flow cytometry analyses were conducted to assess intracellular IL-17 staining in Vδ2 T cells sourced from peripheral blood samples of both HCs and AGA patients. The right panels show the percentage of positively stained cells among Vδ2 T cells. Data of HCs (n=7) and AGA (n=9) are shown. ( B ) Statistical graph showed the levels of various types of cytokines in gout SF (n=6), serum from patients with AGA (n=7) and serum from HCs (n=7). ( C and D ) Flow cytometry analyses of the intracellular staining of IL-17 in Vδ2 T cells ( C ) and in Th17 cells ( D ) of PBMC and SFMC from patients with gout were performed. The right panels show the percentage of positively stained cells among the indicated populations. Data of both groups (n=8) are shown. Results are expressed as mean±SEM. ns, no significance; * P <0.05, ** P <0.01, and *** P <0.001 signify significance as determined through by two-tailed unpaired t -test.

Article Snippet: Fluorescein isothiocyanate (FITC)-conjugated anti-human CD14, phycoerythrin (PE)-conjugated anti-human CD16, allophycocyanine (APC)-conjugated anti-human CD192 (CCR2), PE-conjugated anti-human CD294 (CRTH2), APC-conjugated anti-human CD127 (IL-7Rα), peridinin-chlorophyll-protein/cyanine5.5 (PerCP/Cy5.5) – conjugated anti-human CD117 (c-kit), FITC-conjugated anti-human Lineage Cocktail (CD3, CD14, CD19, CD20, CD56), PerCP/Cy5.5-conjugated anti-human CD3, APC-conjugated anti-human CD57, PerCP-conjugated anti-human CD8, PE/Cy7-conjugated anti-human CD56 (NCAM), FITC-conjugated anti-human CD3, PE-conjugated anti-human TCR γ/δ, APC anti-human TCR Vδ2, PerCP/Cy5.5-conjugated anti-human CD3, FITC-conjugated anti-human TCR Vδ2, APC-conjugated anti-human IL-1β, APC-conjugated anti-human IL-17, APC-conjugated anti-human tumor necrosis factor (TNF) -α, APC-conjugated anti-human interferon (IFN) -γ, PE/Cy7-conjugated anti-human C-X-C chemokine receptor 3 (CXCR3), and APC/Fire 750-conjugated anti-human C-C chemokine receptor 5 (CCR5) antibodies were purchased from BD Biosciences (Franklin Lakes, NJ, USA).

Techniques: Flow Cytometry, Staining, Two Tailed Test

Journal: Journal of Neuroinflammation

Article Title: Infiltrating myeloid cell-derived properdin markedly promotes microglia-mediated neuroinflammation after ischemic stroke

doi: 10.1186/s12974-023-02946-z

Figure Lengend Snippet: Properdin knockout ameliorates microglial activation and inflammation in tMCAO mice. a qPCR assessment of differentially expressed inflammatory genes, including Il1a , Il1b , Il4 , Il6 , Il10 , Nos2 , Ccl2 and Tnf, in the ischemic penumbra 1 d and 3 d after tMCAO. n = 5 mice per group. * P < 0.05 versus the sham group, ** P < 0.01 versus the sham group, # P < 0.05 versus the WT group, ## P < 0.01 versus the WT group, one-way ANOVA with Bonferroni post hoc test. b Immunostaining for TMEM119 (green) and Iba1 (red) and 3D-reconstructed images of TMEM119 + Iba1 + microglia 1 d and 3 d after tMCAO. Scale bar: 50 µm. n = 5 mice per group. * P < 0.05 versus the WT group, ** P < 0.01 versus the WT group, unpaired Student’s t test. c Quantification of Iba1 MFI. d Quantification of average branch length, branch numbers and junctions of 3D-reconstructed microglia in the peri-infarct region. e One day after tMCAO, microglia sorted based on the CD45 int and CD11b + strategy were subjected to PCR array analysis. f FACS analysis of IL-1β, TNF-α and IL-6 expression in microglia 1 d after tMCAO. MFI was quantified. n = 5 mice per group. *** P < 0.001 versus the sham group, ## P < 0.01 versus the WT group, one-way ANOVA with Bonferroni post hoc test

Article Snippet: To evaluate the levels of inflammatory factors in microglia in vivo and in vitro, the cells were further incubated with the following antibodies: PE-anti-TNF-α (Rat, 1:200, Thermo Fisher Scientific, OH, USA, Cat# 12-7321-81, RRID:AB_466198), PE-anti-IL-6 (Rat, 1:200, BioLegend, CA, USA, Cat# 504503, RRID:AB_315337) and APC-anti-IL-1β. (Rabbit, 1:500, Cell Signaling Technology, MA, USA, Cat# 31202, RRID:AB_2799001) All the samples were analyzed by a BD LSRFortessa flow cytometer (BD, NJ, USA), and the data were further analyzed using FlowJo software (Ashland, OR, USA, RRID:SCR_008520).

Techniques: Knock-Out, Activation Assay, Immunostaining, Expressing

Journal: Journal of Neuroinflammation

Article Title: Infiltrating myeloid cell-derived properdin markedly promotes microglia-mediated neuroinflammation after ischemic stroke

doi: 10.1186/s12974-023-02946-z

Figure Lengend Snippet: rmProperdin activates microglia and induces microglia-potentiated neuronal death in vitro. a The mRNA expression of inflammatory factors was measured in primary microglia treated with 2 µg/ml, 4 µg/ml and 8 µg/ml rmProperdin versus control microglia. n = 3 per group, * P < 0.05 versus the control group, ** P < 0.01 versus the control group, *** P < 0.001 versus the control group, n.s, no significance ( P > 0.05), one-way ANOVA with Bonferroni post hoc test. b FACS analysis of IL-1β, TNF-α and IL-6 expression in primary microglia treated with rmProperdin for 24 h. MFI was quantified. n = 3 per group, ** P < 0.01 versus the control group, *** P < 0.001 versus the control group, one-way ANOVA with Bonferroni post hoc test. c Experimental design for the analysis of neuronal viability after treatment with rmProperdin or CM from rmProperdin-treated microglia. d Neuronal viability was assessed by CCK8 assay. n = 6 per group, *** P < 0.001 versus the control group, one-way ANOVA with Bonferroni post hoc test. e Analysis of neuronal death using calcein-AM (green)/PI (red) double staining. Scale bar: 50 µm. f Calcein-AM-positive primary cortical neurons were quantified as percentages of total cells. n = 5 per group, ** P < 0.01 versus the control group, *** P < 0.001 versus the control group, one-way ANOVA with Bonferroni post hoc test

Article Snippet: To evaluate the levels of inflammatory factors in microglia in vivo and in vitro, the cells were further incubated with the following antibodies: PE-anti-TNF-α (Rat, 1:200, Thermo Fisher Scientific, OH, USA, Cat# 12-7321-81, RRID:AB_466198), PE-anti-IL-6 (Rat, 1:200, BioLegend, CA, USA, Cat# 504503, RRID:AB_315337) and APC-anti-IL-1β. (Rabbit, 1:500, Cell Signaling Technology, MA, USA, Cat# 31202, RRID:AB_2799001) All the samples were analyzed by a BD LSRFortessa flow cytometer (BD, NJ, USA), and the data were further analyzed using FlowJo software (Ashland, OR, USA, RRID:SCR_008520).

Techniques: In Vitro, Expressing, Control, CCK-8 Assay, Double Staining