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Anti ccr8

Manufactured by R&D Systems
Sourced in United States

Anti-CCR8 is a laboratory research tool used to detect and study the expression of the CCR8 chemokine receptor. CCR8 is involved in immune cell trafficking and inflammation. This product can be used to identify and quantify CCR8-expressing cells in various biological samples.

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2 protocols using anti ccr8

1

Multiparametric Flow Cytometry Profiling of T Cell Subsets

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Peripheral blood mononuclear cells (PBMCs) were isolated using Ficoll-Plaque Plus density gradient centrifugation (Amersham Biosciences, NJ, USA). Fresh tissues were washed and cut into small fragments, prior to homogenization by tissue disaggregation vessels (BD Medimachine System). The cells in each sample were adjusted to a concentration of 2 × 106 cells/mL, and 0.5 mL cell suspension stimulated with 2μL Leukocyte Activation Cocktail and BD GolgiplugTM (BD Pharmingen, San Diego, CA, USA) for 4 hours.
T cell subsets were phenotyped in isolated PBMCs or tissue cells by flow cytometry (FACSAria flow cytometer, BD Biosciences, NJ, USA) according to manufacturer’s instructions. The cells were labelled with specific monoclonal antibodies; including anti-CD4-PerCP, anti-CD25-PE, anti- Foxp3-FITC, anti-IL-4-PE and anti-IFN-γ-FITC (all from BD Biosciences, NJ, USA). T cell subsets were selected for detailed phenotypic analysis as follow: (1) Th1 cells: IFN-γ+IL-4- CD4+ T cells; (2) Th2 cells: IFN-γ-IL-4+ CD4+T cells; (3) Treg cells: CD4+CD25+Foxp3+ T cells. A minimum of 106 cells per staining were assessed, with at least 105 events being measured. To measure CCRs in peripheral Treg cells, anti-CCR3, anti-CCR4, anti-CCR5 and anti-CCR8 (R & D Systems, Mineapolis, MN, USA) were used to evaluate corresponding receptor expression in circulating Treg cells of 3 groups.
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2

Chemokine Receptor Interaction Assay

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Chemokine (C-C motif) ligand (CCL) 1 (CCL1), CCL2 and chemokine (C-X-C motif) ligand 12 (CXCL12) were purchased from Protein Foundry (Milwaukee, WI). Antibodies were obtained from Abcam (Cambridge, United Kingdom): anti-α1B-AR (host: rabbit; catalog: ab169523), anti-α1D-AR (host: rabbit; catalog: ab84402), anti-CXCR4 (host: goat, ab1670), anti-HA (host: rabbit; catalog: ab9110) ; LifeSpan Biosciences (Seattle, WA): anti-CCR8 (host: goat; catalog: LS-C187704); R&D Systems (Minneapolis, MN): anti-CCR1 (host: mouse; catalog: MAB145), anti-CCR2 (host: mouse; catalog: MAB48607), allophycocyanin (APC) conjugated anti-mouse CD45 (host: rat; catalog: FAB114A), APC conjugated Immunoglobulin G 2B (IgG2B) isotype control (host: rat; catalog: ICO13A), Immunoglobulin G (IgG) isotype control (host: rabbit; catalog: MAB150), IgG isotype control (host: mouse; catalog: MAB004), and IgG isotype control (host: goat; catalog: AB-108-C); and Sigma-Aldrich (St. Louis, MO): anti- FLAG (host: mouse; catalog: F3165).
Phenylephrine (PE), phentolamine (PT), norepinephrine (NE), lipopolysaccharide (LPS) from Pseudomonas aeruginosa (serotype 10.22; source strain ATCC 27316) and all proximity ligation assay (PLA) reagents were purchased from Sigma-Aldrich.
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