Journal: Journal of Investigative Dermatology

Article Title: Differential Production of Th1- and Th2-Type Chemokines by Mouse Langerhans Cells and Splenic Dendritic Cells

doi: 10.1111/j.0022-202x.2004.23607.x

Figure Lengend Snippet: Figure 1 RT-PCR analysis of CXCL10, CXCL9, and CXCL11 gene expression in freshly isolated Langerhans cells (fLC), 48 h cultured LC (cLC), interferon-c (IFN-c)-stimulated cLC (c-cLC), freshly isolated splenic dendritic cells (fDC), 48 h cultured splenic DC (cDC), and IFN-c- stimulated cDC (c-cDC). LC and splenic DC were purified and cul- tured for 48 h in the absence or presence of 100 ng per mL of IFN-g. Samples from fresh and cultured LC and splenic DC were collected and mRNA expression was analyzed using specific primers for each chemokine. CXCL10, CXCL9, and CXCL11 mRNA expression was hardly detectable in fLC. In cLC, mRNA expression of CXCL10 and CXCL11, but not CXCL9, was induced. CXCL9 mRNA was strongly expressed only in g-cLC. In addition, mRNA for CXCL10 and CXCL11 was also strongly expressed in g-cLC. In splenic DC, mRNA for these T helper 1(Th1)-type chemokines was almost undetectable both in fDC and cDC. When stimulated with IFN-g, CXCL10, CXCL9, and CXCL11 mRNA expression was induced. Data are representative of three inde- pendent experiments.

Article Snippet: Measurement of CXCL9, CXCL11, and CCL17 Culture supernatants were collected, stored at 201C, and subjected to the quantification of protein levels of CXCL9, CXCL11, and CCL17 by ELISA using commercially available mouse CXCL9, CXCL11, and CCL17 immunoassay kits (R&D Systems), respectively, according to the manufacturer’s instructions.

Techniques: Reverse Transcription Polymerase Chain Reaction, Gene Expression, Isolation, Cell Culture, Expressing

Journal: Journal of Investigative Dermatology

Article Title: Differential Production of Th1- and Th2-Type Chemokines by Mouse Langerhans Cells and Splenic Dendritic Cells

doi: 10.1111/j.0022-202x.2004.23607.x

Figure Lengend Snippet: Figure 2 Production of T helper 1-type chemokines during culture of Langerhans cells (LC) and splenic dendritic cells (DC). Purified LC and splenic DC were cultured with or without interferon-g (IFN-g), and the concentration of CXCL10, CXCL9, and CXCL11 was measured at different time points (0, 12, 24, 36, 48 h) in the supernatants by ELISA. Representative data of three independent experiments.

Article Snippet: Measurement of CXCL9, CXCL11, and CCL17 Culture supernatants were collected, stored at 201C, and subjected to the quantification of protein levels of CXCL9, CXCL11, and CCL17 by ELISA using commercially available mouse CXCL9, CXCL11, and CCL17 immunoassay kits (R&D Systems), respectively, according to the manufacturer’s instructions.

Techniques: Cell Culture, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Journal of Investigative Dermatology

Article Title: Differential Production of Th1- and Th2-Type Chemokines by Mouse Langerhans Cells and Splenic Dendritic Cells

doi: 10.1111/j.0022-202x.2004.23607.x

Figure Lengend Snippet: Figure 3 Chemotaxis of mCXCR3-transfected 2B4 T cells. Langerhans cells (LC) and splenic dendritic cells (DC) were cultured for 48 h with 100 ng per mL of interferon-g (IFN-g) and the supernatants were collected. The culture supernatants were preincubated with or without neutralizing anti-chemokine monoclonal antibody (mAb) indicated in the figure for 30 min, and assessed for chemotactic activity to CXCR3 transfectant. RPMI 10 medium alone served as a negative control. RPMI 10 medium containing recombinant chemokine served as a positive control. The supernatants of IFN-g-stimulated LC and splenic DC exhibited chemo- tactic activity to CXCR3 transfectant, which is mediated at least by CXCL10 and CXCL9. Mean (SD) (n ¼ 3). Data are representative of three independent experiments.

Article Snippet: Measurement of CXCL9, CXCL11, and CCL17 Culture supernatants were collected, stored at 201C, and subjected to the quantification of protein levels of CXCL9, CXCL11, and CCL17 by ELISA using commercially available mouse CXCL9, CXCL11, and CCL17 immunoassay kits (R&D Systems), respectively, according to the manufacturer’s instructions.

Techniques: Chemotaxis Assay, Transfection, Cell Culture, Activity Assay, Negative Control, Recombinant, Positive Control

Journal: Journal of Investigative Dermatology

Article Title: Differential Production of Th1- and Th2-Type Chemokines by Mouse Langerhans Cells and Splenic Dendritic Cells

doi: 10.1111/j.0022-202x.2004.23607.x

Figure Lengend Snippet: Figure 7 Regulation of T helper 1-type chemokines produced by La- ngerhans cells (LC) and splenic dendritic cells (DC). LC (&) and splenic DC (’) were purified and cultured (1.5 106 cells per mL per 200 mL in each well) for 48 h with or without various stimuli. The supe- rnatants were collected and the concentration of CXCL10 (a), CXCL9 (b), and CXCL11 (c) was measured by ELISA. CXCL10 production by LC was induced by interferon-g (IFN-g), interleukin (IL)-12, lipopoly- saccharide (LPS), Staphylococcus aureus Cowen 1 (SAC), and poly- inosinic–polycytidylic acid (Poly(I:C)). In the case of CXCL9 and CXCL11, only IFN-g induced their production by LC. In the case of splenic DC, the production of CXCL10, CXCL9, and CXCL11 was in- duced by IFN-g, IL-18, LPS, and Poly(I:C). Results are the mean (SD) (n ¼ 4). Significant increase (po0.05) compared with the unstimulated group. Data are representative of four independent experiments.

Article Snippet: Measurement of CXCL9, CXCL11, and CCL17 Culture supernatants were collected, stored at 201C, and subjected to the quantification of protein levels of CXCL9, CXCL11, and CCL17 by ELISA using commercially available mouse CXCL9, CXCL11, and CCL17 immunoassay kits (R&D Systems), respectively, according to the manufacturer’s instructions.

Techniques: Produced, Cell Culture, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: CD11c+ Tbet+ B cells constrain obesity- and vaccination-induced germinal center B cells and T helper cells

doi: 10.1101/2025.09.01.673552

Figure Lengend Snippet: Representative flow cytometry contour plots of PD-1 x CXCR5 expression within CD25-CD4+ T cells isolated from NCD (left) and HFD (right) mouse spleen (top) and liver (bottom) ( A ). Blue gates represent PD-1 HI CXCR5+ cells, yellow gates represent PD-1 HI CXCR5-cells, and black gates represent PD-1-cells ( A ). Frequency and absolute number of PD-1 HI CXCR5+ T FH cells ( B , blue), or PD-1 HI CXCR5-T PH cells ( C , yellow) within the CD25-CD4+ T cell compartments of spleen (top) and liver (bottom) of NCD (grey) and HFD (blue/yellow) mice. Quantification of Bcl-6 ( D ), ICOS ( E ), and T-bet ( F ) protein expression (gMFI) in splenic (left) or hepatic (right) CD25-CD4+ T cell subsets. Representative flow cytometry contour plots depicting relative expression of IL-21 and IFNg by PD-1 HI CXCR5+ T FH cells or PD-1 HI CXCR5-T PH cells from spleen (top) or liver (bottom) that are also CXCR3+ (blue/red) or CXCR3-(black) ( G ). Summary of frequency of IL-21 ( H,J; left) or IFNg ( I,K ; right) producing CD4+ PD-1 HI CXCR5+ T FH (blue) or PD-1 HI CXCR5-T PH (red) cells categorized by CXCR3+ expression as noted in spleen ( H, I ) or liver ( J,K ) from HFD-fed mice. Data pooled from 2 independent experiments with 4 female mice/group, Bar ± SEM; [ Student’s t-test (two-tailed) (B,C); one-way ANOVA (D-F; H-K)]; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001

Article Snippet: C57BL/6J WT female mice were injected IP with 0.25 mg of anti-mouse CXCR3 (CD183) and anti-mouse CXCL9 (MIG) or hamster IgG (BioXCell) every other day for 14 days after completing 12 weeks on HFD.

Techniques: Flow Cytometry, Expressing, Isolation, Two Tailed Test

Journal: Carcinogenesis

Article Title: Chemokine receptor CXCR3 promotes growth of glioma

doi: 10.1093/carcin/bgq224

Figure Lengend Snippet: CXCL9 and CXCL10 are expressed in GL261 glioma cells and/or tumors. (A) RT–PCR identified CXCL10 mRNA in GL261 cells in vitro. GAPDH was used as a control. (B) CXCL10 ELISA showing CXCL10 protein secretion by GL261 cells in vitro at 24 and 48 h (C) CXCL9 and CXCL10 are expressed in intracranial GL261 tumors in vivo as determined by in situ hybridization analysis. Two representative sections, depicting expression of each chemokine, are shown. The colour version of this figure can be found at www.carcin.oxfordjournals.org.

Article Snippet: Recombinant mouse and human CXCL9 and CXCL10 were purchased from R&D Systems.

Techniques: Reverse Transcription Polymerase Chain Reaction, In Vitro, Control, Enzyme-linked Immunosorbent Assay, In Vivo, In Situ Hybridization, Expressing

Journal: Carcinogenesis

Article Title: Chemokine receptor CXCR3 promotes growth of glioma

doi: 10.1093/carcin/bgq224

Figure Lengend Snippet: GL261 tumor-bearing CXCR3-deficient mice have decreased survival rates and tumor-infiltrated NK and NKT cells. (A) Kaplan–Meier survival analysis indicated that CXCR3-deficient mice (n = 10) have shorter life span than WT mice (n = 10, P < 0.0001). Filled squares: CXCR3-deficient mice; filled diamonds: WT mice. (B) Numbers of tumor-infiltrated CD4+, CD8+, Foxp3+, Ly49G2+ and CD11b+ cells were evaluated by immunohistochemistry. Gliomas from CXCR3-deficient mice had a significant reduction of Ly49G2+ (NK and NKT) cells in the tumor as compared with WT mice (**P < 0.01). WT: wild-type; KO: CXCR3-deficient mice. (C) Intratumoral expression of CXCL9 and CXCL10 mRNA was not altered by host CXCR3 deficiency. Shown are representative sections from WT and CXCR3-deficient glioma-bearing mice subjected to in situ hybridization analysis. The colour version of this figure can be found at www.carcin.oxfordjournals.org.

Article Snippet: Recombinant mouse and human CXCL9 and CXCL10 were purchased from R&D Systems.

Techniques: Immunohistochemistry, Expressing, In Situ Hybridization

Journal: Carcinogenesis

Article Title: Chemokine receptor CXCR3 promotes growth of glioma

doi: 10.1093/carcin/bgq224

Figure Lengend Snippet: NBI-74330 suppresses tumor growth in both WT and CXCR3-deficient mice. (A) Kaplan–Meier survival analysis of glioma-bearing WT mice shows that NBI-74330 prolonged animal survival (n = 8), as compared with vehicle-treated mice (n = 8) (P = 0.0212). Filled squares: NBI-74330 treated; filled diamonds: vehicle treated. (B) Kaplan–Meier survival analysis shows that glioma-bearing CXCR3-deficient mice treated with NBI-74330 (n = 7) had a higher survival rate than vehicle-treated glioma-bearing CXCR3-deficient mice (n = 6, P = 0.0028). Filled squares: NBI-74330 treated; filled diamonds: vehicle treated. (C) Similar numbers of tumor-infiltrated lymphocytes and microglia in GL261 gliomas from NBI-74330- and vehicle-treated WT mice. Numbers of tumor-infiltrated CD4+, CD8+, Foxp3+, Ly49G2+ and CD11b+ cells were not affected by NBI-74330 when compared with vehicle treatment. (D) In vivo expression of CXCL9 and CXCL10 was not altered by NBI-74330 treatment. Shown are representative sections from vehicle- and NBI-74330-treated glioma-bearing mice subjected to in situ hybridization analysis. The colour version of this figure can be found at www.carcin.oxfordjournals.org.

Article Snippet: Recombinant mouse and human CXCL9 and CXCL10 were purchased from R&D Systems.

Techniques: In Vivo, Expressing, In Situ Hybridization

Journal: Carcinogenesis

Article Title: Chemokine receptor CXCR3 promotes growth of glioma

doi: 10.1093/carcin/bgq224

Figure Lengend Snippet: NBI-74330 inhibits CXCL9 and CXCL10 stimulation of GS growth or survival in vitro. (A) GL261- and U87-GS cells (2000 cells/ml) were incubated with 1 nM CXCL9 (open squares) or 1 nM CXCL10 (open triangles); U118-, U138-GS cells (5000 cells/ml) and GBM L0 (1000 cells/ml) were incubated with 10 nM CXCL9 (open squares) or 10 nM CXCL10 (open triangles). The control group was cultured in medium without chemokines or growth factors (filled circles). All conditions contained 0.1% dimethyl sulfoxide (NBI-74330 vehicle). Representative results of three individual experiments performed in triplicate are shown. CXCL9 and CXCL10 significantly enhanced GL261- and U87-GS growth at day 6 and 9 (*P < 0.05, **P < 0.01) and prevented U138-GS cell loss at day 9. CXCL9 stimulation significantly increased cell numbers of GBM L0 at day 9 (#P < 0.05); CXCL10-stimulated group was not statistically significant as compared with control. (B) The effect of 1 μM NBI-74330 (black filled bars) and 0.1% dimethyl sulfoxide (open bars) on chemokine- and growth factor-stimulated GS growth. Representative results of three individual experiments performed in triplicate are shown. NBI-74330 attenuated response of GL261-, U87- and U138-GS to CXCL9 and CXCL10 but did not affect either control or growth factor-supplemented groups.

Article Snippet: Recombinant mouse and human CXCL9 and CXCL10 were purchased from R&D Systems.

Techniques: In Vitro, Incubation, Control, Cell Culture

Journal: Inflammation

Article Title: Programmed death-ligand 1 (PD-L1) Modulates Chemokine Production Via the TLR4/TRAF6 Signaling Axis During LPS + IFN-γ-Induced Endotoxemia-mimicked Sepsis

doi: 10.1007/s10753-025-02394-2

Figure Lengend Snippet: PD-L1 mediates the regulation of chemokine biosynthesis in M1-polarized macrophages, as determined by RNA-seq. ( A ) PD-L1 NC /THP-1-M and PD-L1 HI /THP-1-M cells were treated with LPS (100 ng/mL) + IFN-γ (20 ng/mL) for 6 h and subjected to RNA sequencing analysis. The KEGG database was used to identify enriched signaling pathways whose activation significantly differed (up). Single-gene GSEA was conducted using the GSE57065 , GSE65682 , and GSE95233 datasets to identify the relationship between PD-L1 and chemokine signaling pathways (down). ( B ) Intersection analysis of upregulated genes between the PD-L1 HI /THP-1-M + LPS + IFN-γ group and PD-L1 NC /THP-1-M + LPS + IFN-γ group and between the PD-L1 NC /THP-1-M + LPS + IFN-γ group and PD-L1 NC /THP-1-M + PBS group via a Venn diagram, followed by further intersection with chemokine datasets, revealed three highly expressed PD-L1-mediated chemokines (CCL8, CXCL9, and CXCL10) under LPS + IFN-γ stimulation. ( C ) RNA sequencing analysis revealed the expression of CCL8, CXCL9 and CXCL10 upon PD-L1 overexpression. ( D ) PD-L1 NC /THP-1-M and PD-L1 HI /THP-1-M cells were treated with LPS + IFN-γ for 12 h, after which CCL8 and CXCL9 mRNA and protein levels were quantified ( n = 3 independent biological replicates). ( E ) Human monocytes were induced into macrophages, transfected with scramble siRNA and PD-L1 siRNA, and then treated with LPS + IFN-γ for 12 h. CCL8 and CXCL9 mRNA and protein levels were quantified ( n = 3 independent biological replicates). ( F ) PD-L1 NC /THP-1-M and PD-L1 KO /THP-1-M cells were treated with LPS + IFN-γ for 12 h, after which CCL8 and CXCL9 mRNA and protein levels were quantified ( n = 3 independent biological replicates). The data are presented as the mean ± SEM. Statistical analysis for D, E and F was performed by one-way ANOVA with LSD test for post hoc analyses. * p < 0.05

Article Snippet: CCL8 and CXCL9 concentrations in cell supernatants or plasma were quantified using ELISA kits (Boster Biological Technology; Wuhan, China) with absorbance measurements at 450 nm, following the manufacturer’s protocols.

Techniques: RNA Sequencing, Protein-Protein interactions, Activation Assay, Expressing, Over Expression, Transfection

Journal: Inflammation

Article Title: Programmed death-ligand 1 (PD-L1) Modulates Chemokine Production Via the TLR4/TRAF6 Signaling Axis During LPS + IFN-γ-Induced Endotoxemia-mimicked Sepsis

doi: 10.1007/s10753-025-02394-2

Figure Lengend Snippet: TLR4 acts as a critical upstream regulator of PD-L1 in modulating CCL8 and CXCL9 expression. ( A ) GSEA of RNA-seq data to investigate the relationship between PD-L1 expression and Toll-like receptor signaling pathway activation. ( B ) PD-L1 HI /THP-1-M and PD-L1 NC /THP-1-M cells were transfected with scramble siRNA or TLR4 siRNA, respectively, followed by 12 h of stimulation with LPS (100 ng/mL) + IFN-γ (20 ng/mL). The supernatants and cells were harvested for concurrent measurements of CCL8 and CXCL9 protein and mRNA levels via ELISAs and qRT‒PCR ( n =3 independent biological replicates). The data are presented as the mean ± SEM. Statistical analysis for B was performed by one-way ANOVA with LSD test for post hoc analyses. * p < 0.05

Article Snippet: CCL8 and CXCL9 concentrations in cell supernatants or plasma were quantified using ELISA kits (Boster Biological Technology; Wuhan, China) with absorbance measurements at 450 nm, following the manufacturer’s protocols.

Techniques: Expressing, RNA Sequencing, Activation Assay, Transfection

Journal: Inflammation

Article Title: Programmed death-ligand 1 (PD-L1) Modulates Chemokine Production Via the TLR4/TRAF6 Signaling Axis During LPS + IFN-γ-Induced Endotoxemia-mimicked Sepsis

doi: 10.1007/s10753-025-02394-2

Figure Lengend Snippet: PD-L1 binds to the TLR4 downstream molecule TRAF6 to regulate CCL8 and CXCL9 expression. ( A ) Venn diagram analysis of proteins coimmunoprecipitated with PD-L1 in THP-1-M cells stimulated with LPS (100 ng/mL) + IFN-γ (20 ng/mL) for 6 h via IP‒MS. After excluding proteins bound to the IgG control group, 289 PD-L1-specific interacting proteins were identified (up). Representative MS/MS spectrum showing peptide fragments of the TRAF6 protein identified in the immunoprecipitation complex (down). ( B ) THP-1-M (up) and human monocyte-derived (down) macrophages were treated with or without LPS+IFN-γ for 6 h and then harvested for Co-IP. TRAF6 protein levels were measured via Western blotting. ( C ) PD-L1 HI /THP-1-M and PD-L1 NC /THP-1-M cells were transfected with scramble siRNA or TRAF6 siRNA, respectively, followed by 12 hours of stimulation with LPS+IFN-γ. The supernatants and cells were harvested for concurrent measurements of CCL8 and CXCL9 protein and mRNA levels via ELISAs and qRT‒PCR ( n =3 independent biological replicates). ( D ) Structural models of full-length PD-L1 and TRAF6 predicted by AlphaFold (PAE plots were obtained from AlphaFold). ( E ) Molecular docking generic matching information. ( F ) A protein‒protein interaction cartoon model generated by PyMOL (left). Predicted binding sites between PD-L1 and TRAF6 (right). ( G ) PD-L1 NC /THP-1-M, PD-L1 HI /THP-1-M and PD-L1 Δ62-139 /THP-1-M cells were treated with LPS+IFN-γ for 12 h. The supernatants and cells were harvested for concurrent measurements of CCL8 and CXCL9 protein and mRNA levels via ELISAs and qRT‒PCR ( n =3 independent biological replicates). The data are presented as the mean ± SEM. Statistical analysis for C and G was performed by one-way ANOVA with LSD test for post hoc analyses. * p <0.05.

Article Snippet: CCL8 and CXCL9 concentrations in cell supernatants or plasma were quantified using ELISA kits (Boster Biological Technology; Wuhan, China) with absorbance measurements at 450 nm, following the manufacturer’s protocols.

Techniques: Expressing, Control, Tandem Mass Spectroscopy, Immunoprecipitation, Derivative Assay, Co-Immunoprecipitation Assay, Western Blot, Transfection, Generated, Binding Assay

Journal: Inflammation

Article Title: Programmed death-ligand 1 (PD-L1) Modulates Chemokine Production Via the TLR4/TRAF6 Signaling Axis During LPS + IFN-γ-Induced Endotoxemia-mimicked Sepsis

doi: 10.1007/s10753-025-02394-2

Figure Lengend Snippet: PD-L1 knockout attenuates septic mouse mortality and suppresses CCL8 and CXCL9 expression. ( A – C ) PD-L1 WT /mice and PD-L1 KO /mice were intraperitoneally injected with LPS (15 mg/kg) and IFN-γ (10 µg/kg) to induce endotoxemia-induced sepsis. ( B ) The survival rate was determined over a 60-hour observation period and is presented as a Kaplan‒Meier survival curve ( n = 15 mice per group). The results are presented as Kaplan–Meier survival curves. The results of the statistical analysis were analyzed with the log-rank test. * p < 0.05. ( C ) Peripheral blood was collected 12 h posttreatment for serum isolation and PBMC separation. CCL8 and CXCL9 protein levels in serum and their corresponding mRNA levels in PBMCs were subsequently analyzed ( n = 3 independent biological replicates). ( D ) Peritoneal macrophages (PMs) were extracted from PD-L1 wild-type and knockout mice and then treated with LPS (100 ng/mL) + IFN-γ (20 ng/mL) for 12 h. The cells were harvested for CCL8 and CXCL9 mRNA and protein detection by qRT‒PCR and ELISAs ( n = 3 independent biological replicates). (E) Peripheral blood monocytes were extracted from PD-L1 wild-type and knockout mice and then treated with LPS + IFN-γ for 12 h. The cells were harvested for CCL8 and CXCL9 mRNA and protein detection by qRT‒PCR and ELISAs ( n = 3 independent biological replicates). The data are presented as the mean ± SEM. Statistical analysis for C was performed by Student’s t test, and that for D and E was performed by one-way ANOVA with LSD test for post hoc analyses. * p < 0.05

Article Snippet: CCL8 and CXCL9 concentrations in cell supernatants or plasma were quantified using ELISA kits (Boster Biological Technology; Wuhan, China) with absorbance measurements at 450 nm, following the manufacturer’s protocols.

Techniques: Knock-Out, Expressing, Injection, Isolation

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Evaluation of the 5-day assay precision.

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques: Concentration Assay, Control, Clinical Proteomics

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Dilution linearity. Linear regression analysis ( n = 3) of the HISCL CXCL9 immunoassay using serum and plasma samples. R 2 , coefficient of determination.

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques: Clinical Proteomics

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Spike-recovery test.

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques: Clinical Proteomics

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Cross-reactivity with other chemokines, compounds, and drugs. We observed no cross-reactivity with structurally similar chemokines (10 ng/mL; top). No interference was observed between CXCL9 and haemoglobin (46.0 g/L), conjugated bilirubin (BIL-F; 3450 µmol/L), unconjugated bilirubin (BIL-C; 3540 µmol/L), chyle (17,700 FTU), purified RF (500 IU/mL), and 100 μg/mL of various drugs (bottom).

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques: Purification

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Frequency distribution of serum CXCL9 levels in healthy individuals. Results are shown as individual numbers of CXCL9 levels (open bars). The dashed line indicates the 95 percentile: 39.0 pg/mL.

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques:

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Distribution of serum CXCL9 levels. Levels of CXCL9 in HCs, COVID-19 patients not admitted to the ICU (Non-ICU) and those admitted to the ICU (ICU), and patients with asthma, ILDs, and HP, respectively. Results are shown as individual data points (circles) with medians (bars) and interquartile ranges (box). Whiskers extend to the minimum and maximum values excluding outliers.

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques:

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Age-related differences in CXCL9 levels in patients with asthma. Distributions of serum ( a ) CXCL9, ( b ) CXCL10, ( c ) CXCL11, ( d ) IL-4, and ( e ) total IgE levels according to age, which is indicated on the x-axis. Results are shown as individual data points with medians (bars) and interquartile ranges (box). Whiskers extend to the minimum and maximum values excluding outliers. The ratios of ( f ) atopic asthma, ( g ) comorbidity of AD (atopic dermatitis), ( h ) comorbidity of other atopic diseases (AD, atopic rhinitis, atopic conjunctivitis), ( i ) childhood-onset asthma, ( j ) medication steps in JGL2015 (Blue: I, Green: II, Yellow: III, Red: IV) and ( k ) sex are indicated. Filled bars indicate positivity or female for each condition. P-values were calculated using the Steel–Dwass test. *P < 0.05, **P < 0.01, ***P < 0.001, Mann–Whitney U (†) ( a – d ), and Fisher's exact tests ( f – k ).

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques: MANN-WHITNEY

Journal: Scientific Reports

Article Title: Development of a new HISCL automated CXCL9 immunoassay

doi: 10.1038/s41598-023-32513-8

Figure Lengend Snippet: Correlation of biomarkers with CXCL9 levels in patients with asthma.

Article Snippet: Whereas to determine the LoQ, CXCL9 recombinant protein (R&D systems, 392-MG, lot ABU1614052) was diluted with sample buffer (0.72–13.8 pg/mL), and samples and dilutions were tested over 3 days (1 run/day) on one instrument (lots A and B; n = 30/sample).

Techniques:

Journal: bioRxiv

Article Title: An Inflammatory Clock Predicts Multi-morbidity, Immunosenescence and Cardiovascular Aging in Humans

doi: 10.1101/840363

Figure Lengend Snippet: Decomposition of the inflammatory score was conducted by estimating the most variable jacobians (first order partial derivative of inflammatory age) ( A ). Both positive and negative contributors to inflammatory age are observed. The top 15 most variable jacobians are CXCL9, EOTAXIN, Mip-1α, LEPTIN, IL-1β, IL-5, IFN-α and IL-4 (positive contributors) and TRAIL, IFN-β, CXCL1, IL-2, TGF-α, PAI-1 and LIF (negative contributors) (A). In a validation study, 97 healthy adults (age 25-90) well matched for cardiovascular risk factors, were selected form a total of 151 recruited subjects. Immune protein analysis was conducted in samples from these subjects. CXCL9, HGF, CXCL1 and LIF were found to change in the same direction in both the Stanford 1KIP and the validation cohort ( A and B ). Cardiovascular age was estimated using 3 parameters: (1) aortic pulse wave velocity, a measure of vascular stiffness; (2) relative wall thickness (RWT), a measure of ventricular remodeling, and (3) early diastolic mitral annular velocities (e’), a measure of ventricular relaxation. After adjusting for age, sex, BMI, heart rate, systolic blood pressure, fasting glucose and total cholesterol to HDL ratio, positive correlations were obtained between CXCL9 and PWV ( R = 0.22) and RWT ( R = 0.3) ( P < 0.05), and negative correlations were observed between LIF and PWV ( R = −0.27), and RWT ( R = −0.22) ( C and D ). No variable included in the models had high co-linearity as suggested by variance inflation factors (VIF) < 3 for each factor. Induced pluripotent stem cells (hiPSCs) were obtained from isolated fibroblasts ( N = 5, in duplicates) using the Yamanaka factors and differentiated them into endothelial cells (hiPSC-ECs) under well-defined conditions as previously described . Expression levels of CXCL9 and SIRT3 were measured by RT-PCR as described under Methods. A significant age-dependent increase in CXCL9 mRNA expression levels is observed ( P < 0.01), which reaches a plateau after the sixth cell passage ( E ). Concomitant with the increase in CXCL9, down-regulation in SIRT3 mRNA can be observed after the second cell passage ( P < 0.01) (E). Addition of increasing doses (10 to 800 ng/ml) of exogenous CXCL9 to young (day 7) hiPSC-ECs induces down-regulation of SIRT3 mRNA expression (F). Expression of the CXCL9 receptor, CXCR3, was measured in young cardiomyocytes derived from hiPSCs (hiPSC-CM) as well as in hiPSC-ECs, HUVEC cells, freshly isolated fibroblasts and hiPSCs. Elevated expression is observed in hiPSC-ECs, HUVEC cells but not in other cell types ( F ) suggesting that the endothelium but no other cell subsets is target of CXCL9 and potentially other CXCR3 ligands as well.

Article Snippet: Following normalization, the vessels were incubated with either PBS or different concentrations of recombinant mouse CXCL9 protein (R&D systems, catalog number 492-MM) for 3-4 hrs.

Techniques: Biomarker Discovery, Isolation, Expressing, Reverse Transcription Polymerase Chain Reaction, Derivative Assay

Journal: bioRxiv

Article Title: An Inflammatory Clock Predicts Multi-morbidity, Immunosenescence and Cardiovascular Aging in Humans

doi: 10.1101/840363

Figure Lengend Snippet: To validate the association between CXCL9 expression and SIRT3 in hiPSC-ECs, CXCL9 was knockdown (KD) in hiPSCs using shRNA. Quantitative PCR data show significant knockdown of CXCL9 in hiPSCs following transfection with CXCL9-shRNA compared to scramble controls ( A ). hiPSC-ECs failed to show an increase in CXCL9 expression in CXCL9-KD cells as they were passaged when compared to scramble controls ( B ). On the contrary, CXCL9-KD hiPSC-ECs show reversal of their SIRT3 levels in serially passaged hiPSC-ECs when compared to scramble controls ( C ). Quantification of the number of capillary-like networks formed by scramble- and CXCL9-KD hiPSC-ECs show that CXCL9-KD hiPSC-ECs retain their capacity to form tubular networks even at later passages when compared to scramble controls that showed impaired tube formation at later passages of hiPSC-ECs ( D ). Line graph of percent relaxation of mouse thoracic aortic sections to Acetylcholine show impaired vascular reactivity to increasing concentrations of CXCL9, suggesting CXCL9 impairs vascular function ( E ). Significance of impaired vascular reactivity was determined by 2-way ANOVA, followed by a Bonferroni post-test, N=3, n=3-4, *p<0.05.

Article Snippet: Following normalization, the vessels were incubated with either PBS or different concentrations of recombinant mouse CXCL9 protein (R&D systems, catalog number 492-MM) for 3-4 hrs.

Techniques: Expressing, Knockdown, shRNA, Real-time Polymerase Chain Reaction, Transfection

Journal: bioRxiv

Article Title: An Inflammatory Clock Predicts Multi-morbidity, Immunosenescence and Cardiovascular Aging in Humans

doi: 10.1101/840363

Figure Lengend Snippet: Representative images of capillary-like networks from scramble- and CXCL9-KD hiPSC-ECs show that CXCL9-KD hiPSC-ECs retain their capacity to form tubes even at later passages when compared to scramble that showed impaired tube formation towards later passages of hiPSC ECs.

Article Snippet: Following normalization, the vessels were incubated with either PBS or different concentrations of recombinant mouse CXCL9 protein (R&D systems, catalog number 492-MM) for 3-4 hrs.

Techniques:

Journal: Diagnostics

Article Title: An Antibody-Aptamer-Hybrid Lateral Flow Assay for Detection of CXCL9 in Antibody-Mediated Rejection after Kidney Transplantation

doi: 10.3390/diagnostics12020308

Figure Lengend Snippet: Summary of used ssDNAs and their characterization (all obtained from ELLA Biotech, Planegg, Germany).

Article Snippet: Commercial CXCL9 (Cat# TP720369) was acquired from OriGene (MD, USA).

Techniques: Sequencing, Modification, Control

Journal: Diagnostics

Article Title: An Antibody-Aptamer-Hybrid Lateral Flow Assay for Detection of CXCL9 in Antibody-Mediated Rejection after Kidney Transplantation

doi: 10.3390/diagnostics12020308

Figure Lengend Snippet: The 1st ( A ) and 2nd patient cohort ( B ): CXCL9 was determined using the Luminex-based multiplex protein array in the cryo-preserved EDTA-processed plasma samples of the kidney transplant recipients at the day of the biopsy. Boxplots show the median value, the mean and the standard error of the mean. Wilcoxon test revealed a statistical significance between groups with * p < 0.05; ** p < 0.01; *** p < 0.001 as indicated. TCMR = T-cell mediated rejection, AMR = antibody-mediated rejection.

Article Snippet: Commercial CXCL9 (Cat# TP720369) was acquired from OriGene (MD, USA).

Techniques: Luminex, Multiplex Assay, Protein Array, Clinical Proteomics

Journal: Diagnostics

Article Title: An Antibody-Aptamer-Hybrid Lateral Flow Assay for Detection of CXCL9 in Antibody-Mediated Rejection after Kidney Transplantation

doi: 10.3390/diagnostics12020308

Figure Lengend Snippet: Multiplex ELISA analysis in biopsy lysates of the 1st patient cohort. Protein lysates were identified as inner cortex using microscopic analysis and grouped into non-rejection ( n = 11), borderline-TCMR ( n = 6), TCMR (T-cell mediated rejection, n = 9) and AMR (antibody-mediated rejection, n = 18). In total, 12.5 µg total protein was used to determine the concentrations of the cytokines/chemokines (( A ) CXCL9, ( B ) CCL5, ( C ) CXCL10 and ( D ) HGF) using Luminex-based multiplex assay. Boxplots show the median value, the mean and the standard error of the mean. Wilcoxon test revealed a statistical significance between groups with * p < 0,05, ** p < 0.01, *** p < 0.001 as indicated.

Article Snippet: Commercial CXCL9 (Cat# TP720369) was acquired from OriGene (MD, USA).

Techniques: Multiplex Assay, Enzyme-linked Immunosorbent Assay, Luminex

Journal: Diagnostics

Article Title: An Antibody-Aptamer-Hybrid Lateral Flow Assay for Detection of CXCL9 in Antibody-Mediated Rejection after Kidney Transplantation

doi: 10.3390/diagnostics12020308

Figure Lengend Snippet: Schematic of the basic structure of the developed aptamer-antibody-hybrid-lateral flow assay (hybrid-LFA) with sample pad, nitrocellulose membrane and absorbent pad, providing the necessary lateral flow fluids and overlapping glued to a backing card. The hybrid-LFA uses a capture antibody in the test zone (TZ) and a detection aptamer conjugated to gold nanoparticles both with high affinity for CXCL9. ( A ) test platform composition, ( B ) positive test (two red lines, TZ and CZ), ( C ) negative test (one red line, CZ) (modified according to Seiler et al. ).

Article Snippet: Commercial CXCL9 (Cat# TP720369) was acquired from OriGene (MD, USA).

Techniques: Lateral Flow Assay, Membrane, Modification

Journal: Diagnostics

Article Title: An Antibody-Aptamer-Hybrid Lateral Flow Assay for Detection of CXCL9 in Antibody-Mediated Rejection after Kidney Transplantation

doi: 10.3390/diagnostics12020308

Figure Lengend Snippet: Lateral Flow Assays using different concentrations of purified CXCL9 in pooled urine samples of patients with unsuspicious finding proven by biopsy and binding buffer (1:1) with the corresponding columns reflecting the signal intensity of the test line. Scans were taken 45 min after sample application. N = 1.

Article Snippet: Commercial CXCL9 (Cat# TP720369) was acquired from OriGene (MD, USA).

Techniques: Purification, Binding Assay