Purified monoclonal anti-mouse CXCR3 (CXCR3–173) and its isotype control hamster IgG used for injection were obtained from BioXCell. For flow cytometry, anti-CD4, anti-CD8, anti-CD44, and anti-CXCR3 antibodies were purchased from BioLegend. For immunohistological chemistry, anti-TNF-α and anti-claudin-1 antibodies were purchased from Abcam. For immunofluorescence staining, anti-aquaporin-5 (AQP5) and Alexa Fluor647-conjugated rabbit IgG were purchased from Abcam.
Anti cxcr3
Manufactured by BioLegend
Sourced in United States
Anti-CXCR3 is a lab equipment product that detects the CXCR3 chemokine receptor. CXCR3 is a G protein-coupled receptor that plays a role in immune cell trafficking and inflammation. The Anti-CXCR3 product can be used to identify and study cells expressing the CXCR3 receptor.
Automatically generated - may contain errors
Lab products found in correlation
Anti ccr6, by BioLegend (3 mentions)
Anti ccr4, by BioLegend (3 mentions)
Anti cd3, by BioLegend (3 mentions)
Anti cd4, by BioLegend (2 mentions)
Anti cd44, by BioLegend (2 mentions)
Anti cd69, by BioLegend (2 mentions)
Anti tigit, by BioLegend (2 mentions)
Anti pd 1, by BioLegend (2 mentions)
Cxcl10, by Thermo Fisher Scientific (2 mentions)
Cxcl9, by Thermo Fisher Scientific (2 mentions)
Anti ccr7, by R&D Systems (2 mentions)
Ccl21, by Thermo Fisher Scientific (2 mentions)
Ccl19, by Thermo Fisher Scientific (2 mentions)
Lsr fortessa cytometer, by BD (2 mentions)
Live dead aqua, by Thermo Fisher Scientific (2 mentions)
Foxp3 fix perm kit, by Thermo Fisher Scientific (2 mentions)
Anti cd38, by BD (2 mentions)
Anti igd, by Thermo Fisher Scientific (2 mentions)
Anti cd27, by Thermo Fisher Scientific (2 mentions)
Unlabeled mouse igg, by BioLegend (2 mentions)
10 protocols using anti cxcr3
1
Quantifying Immune Cell Infiltration and Inflammation
2
Immune Cell Profiling in EAE Mice
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Spinal cords and spleens were separated from naive mice and PBS-, shNC-MSC-, or shBecn1-MSC-treated EAE mice after euthanasia. Mononuclear cells were isolated from spinal cords with Percoll (GE healthcare 17-0891-09) or from spleens with Ficoll (LymphoprepTM, 1114547). Cells were stained with anti-CD4 (eBioscience, 17-0041-83; BD Biosciences, 553729), anti-CD8 (BD Biosciences, 553035), anti-PTPRC (45-0452-82), anti-LY6G (11-5931-82), anti-ITGAM (BD Biosciences, 553312), anti-IL2RA (12-0251-83), anti-CD69 (11-0691-82), anti-CXCR3 (12-1831-80), and anti-CCR6 antibodies (Biolegend, 129814). For Th1 cell, Th17 cell and Treg cell analysis, cells were stained with surface marker, permeabilized with the Intracellular Fixation and Permeabilization Buffer Set (eBioscience, 88-8824-00), and then stained with anti-IFNG (11-7311-82), anti-IL17A (12-7177-81), and anti-FOXP3 (12-5773-80) antibodies. All these antibodies were purchased from eBioscience, unless marked otherwise. The samples were analyzed by flow cytometry (FACSAria II, BD Biosciences, San Jose, CA USA)
3
Transwell Migration Assay for CD8 T Cells
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CD8a microbeads (Miltenyi Biotec) were used to separate CD8 lymphocytes from 6 to 8 weeks old Balb/c mice. They were then stimulated with anti-CD3/CD28 Dynabeads (Thermo Fisher Scientific) at a 1:1 beads/cells ratio and 100 U/mL human IL-2 (PeproTech) for 96 hours. Transwell inserts with a pore size of 5.0 m. 5×105 CD8 T cells were put into the top chamber of the inserts (5.0 µm pore size, Corning). The bottom of the bottom well was filled with IMDM medium or conditioned medium (CM) from CT26 control or GBP2-KO cells, with or without CXCL10/11 overexpression. In order to block CXCR3, CD8 T cells were first incubated with 10 µg/mL of anti-CXCR3 (BioLegend, 126517) for 30 min before being added to the top chamber. Plates were kept at 37°C overnight, and the contents of the lower chamber were taken. Trypan blue was used to count the number of CD8 +T cells that were still alive. Detailed information of other experiments protocols was also described in online supplemental information .
4
Comprehensive NK Cell Phenotyping
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The following fluorescently conjugated antibodies were used for phenotypic analysis of NK cells: anti-LFA-1 (363404); anti-CXCR3 (353720); anti-PD-1 (329906); anti-CD107a (328612); anti-NKP44 (325116); anti-CD158e1 (312706); anti-CD158d (347006); anti-CD27 (356412); anti-CCR4 (359412); anti-DNAM-1 (338316); anti-CD16 (302040); anti-Granzyme B (515406); anti-CD62L (304806); anti-CD69 (310910); anti-NKP80 (346706); anti-NKP30 (325210); anti-CD158f (341304); anti-CD158b (312612); anti-Tim-3 (345012); anti-CD94 (305504); anti-TIGIT (372706); anti-TRAIL (308206); anti-CD57 (322306); anti-CX3CR1 (341610); anti-NKG2D (320808); anti-Perforin (353310); anti-Ki67 (350504); anti-IFN-γ (506518); anti-CD94 (305506); anti-NKp46 (331916); anti-CTLA4 (369614); anti-CD96 (338416); anti-41BB (309818); anti-CD25 (356108) were purchased from Biolegend. anti-CD159a/NKG2A (FAB1059P) and anti-NKG2C (FAB138G) were purchased from R&D.
The following fluorescently conjugated antibodies were used to identify immune cell types in eNK or PBMC: anti-human Lineage Cocktail (348803); anti-CD56 (362550); anti-CD3 (300430); anti-CD33 (366620); anti-HLA-DR (307606); anti-CD14 (301836); anti-CD19 (302242); anti-CD11b (301322); anti-CD25 (356108); and anti-FOXP3 (320108) were purchased from Biolegend, San Diego, CA, USA.
The following fluorescently conjugated antibodies were used to identify immune cell types in eNK or PBMC: anti-human Lineage Cocktail (348803); anti-CD56 (362550); anti-CD3 (300430); anti-CD33 (366620); anti-HLA-DR (307606); anti-CD14 (301836); anti-CD19 (302242); anti-CD11b (301322); anti-CD25 (356108); and anti-FOXP3 (320108) were purchased from Biolegend, San Diego, CA, USA.
5
Transwell Migration of T Cells
6
Comprehensive B Cell Immunophenotyping
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For analysis of B cells from blood, cultures or nasal tissue, cell samples were simultaneously Fc-blocked with unlabeled mouse IgG (Lampire) and ICAM-1-blocked with anti-ICAM-1, (BioLegend), which was fluorescently-labeled or not, depending upon the experiment. After 30 minutes at 4°C, B cells were stained with fluorescently tagged virus (Alexa Fluor 488-RV-A39 and Alexa Fluor 568-RV-A16), viability dye Live/Dead Aqua (ThermoFisher), and various combinations of the following fluorescent antibodies depending on the sample type and application: anti-CD3 (BioLegend), anti-CD11c (BioLegend), anti-CD19 (BioLegend), anti-CD20 (BioLegend), anti-CD27 (ThermoFisher), anti-CD38 (Becton Dickinson), anti-CCR5 (ThermoFisher), anti-CXCR3 (BioLegend), anti-CXCR5 (BioLegend), anti-IgD (ThermoFisher), anti-IgM (BioLegend), anti-IgG (BD Biosciences), anti-IgA (Miltenyi), and anti-IgE (BioLegend). After incubating for 30 minutes at 4°C, cells were then fixed and permeabilized (FoxP3 fix/perm kit, ThermoFisher), before staining for intracellular IgM (BioLegend), IgG (Becton Dickinson), IgA (Miltenyi), IgE (BioLegend), Ki-67 (BioLegend), and T-bet (BioLegend). Cells were analyzed on an LSR Fortessa Cytometer (Becton Dickinson) using FlowJo version 10.5.3 (TreeStar). (See Table S1 ).
7
Comprehensive B Cell Immunophenotyping
8
OSCC Secretome Effects on Teff and Tregs
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Sorted Teff and Tregs from healthy donors were activated with anti-CD3/CD28 beads (1:5 ratio) (Life Technologies) and 1000 UI IL-2 for 5 days a 37°C. Then, 100 uL of OSCC and control secretomes were added to 2x105 Teff or 2x105 Tregs (in 100uL) in XVIVO-15 serum-free medium 48h a 37°C. After the incubation, the supernatants were stored for further cytokine production measurement using the Cytokine Bead Array Th1/2/17 Kit (BD) and the cells were counted (CountBright Absolute Counting Beads), stained with Live/Dead dye (Life Technologies), anti-CXCR3, anti-CCR4, anti-CCR6, anti-CCR8, anti-PD-1 and anti-TIGIT (all BioLegend) and analyzed by flow cytometry. For the analysis of cells after secretome co-culture, cells were washed after co-culture with secretome and cultured in new media X-VIVO15 (LONZA) serum-free medium for 48 h at 37°C with anti-CD3/CD28 beads (1:5 ratio) (Life Technologies) and 1000 UI IL-2. After the incubation, the supernatants were stored for further cytokine production measurement and the cells were stained with Live/Dead dye (Life Technologies), anti-CCR6, anti-CCR8, anti-PD-1 and anti-TIGIT (all BioLegend).
9
Transwell Assay for T Cell Migration
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Migration of T cells was assessed by using Transwell permeable supports with 5-μm polycarbonate membrane (Costar, 3387). To determine cell migration in response to soluble factors, the bottom chamber was loaded with 0.1% FBS, 200 ng/mL CCL21 (PeproTech, 250–13), 200 ng/mL CCL19 (PeproTech, 250–27B), 150 ng/mL CXCL9 (PeproTech 250–18), or 50 ng/mL CXCL10 (PeproTech, 250–16) in T-cell medium. T cells were incubated for 2 (naïve) or 3 hours (activated) at 37°C and migrated cells in the bottom chamber were collected and counted by flow cytometry using Precision Count Beads. According to the experimental requirements, activated T cells were pretreated for 1 hour with 10 μg/mL anti-CCR7 (R&D Systems, 4B12), 250 μg/mL anti-CXCR3 (BioLegend, CXCR3–173), 100 μmol/L Rac1 inhibitor NSC23766, or the appropriate isotype, or vehicle control.
10
Characterization of Murine and Human Immune Cells
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Both OTX015 and JQ1 (Stratech Scientific, UK) were dissolved in DMSO to make a stock solution of 10 mM. Aliquots were kept at -80 o C for up to 6 months and diluted with appropriate culture media for in vitro use. Murine antibodies for cell culture (anti-CD3, anti-CD28, anti-IL-4, anti-IFN-) and flow cytometry (anti-IL-17, anti-IFN-) were purchased from eBioscience (USA) and human antibodies (anti-IL-17, anti-IFN-) were purchased from Biolegend (USA) or eBioscience (USA). Sorting antibodies (anti-CD3, anti-CD4, anti-CD44, anti-CD62L for murine and anti-CD45RA, anti-CD45RO, anti-CCR7, anti-CCR6, anti-CCR4, anti-CXCR3 for human) were purchased from Biolegend (USA).
Recombinant murine IL-6 and IL-12 were purchased from PeproTech (USA) and recombinant human TGF- was purchased from R&D systems (USA).
Recombinant murine IL-6 and IL-12 were purchased from PeproTech (USA) and recombinant human TGF- was purchased from R&D systems (USA).
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