Ccr2 apc

Manufactured by R&D Systems

CCR2-APC is a fluorophore-conjugated antibody that recognizes the CCR2 protein. CCR2 is a chemokine receptor that plays a role in the recruitment of monocytes and macrophages to sites of inflammation. The APC (Allophycocyanin) fluorophore allows for the detection of CCR2-expressing cells by flow cytometry.

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Lab products found in correlation

Fc block, by BD (3 mentions) Efluor 450, by Thermo Fisher Scientific (3 mentions) Alexa fluor 647 protein labeling kit, by Thermo Fisher Scientific (2 mentions) Cd8 percp cy5, by BD (2 mentions) Ly6c percp cy5, by BD (2 mentions) Cd3 apc 145 2c11, by BD (2 mentions) Cd4 apc, by R&D Systems (2 mentions) Cd11c v450 or pe cy7 hl3, by BD (2 mentions) Siglecf bv421 e50 2440, by BD (2 mentions) Cd31 percp efluor 710 390, by R&D Systems (2 mentions) Efluor 450, by R&D Systems (2 mentions) Fasl apc, by R&D Systems (2 mentions) Accucount particles, by Spherotech (2 mentions) F4 80 pe, by BioLegend (1 mentions) Cd45 af700, by BioLegend (1 mentions) Cd80 pe cy7, by BioLegend (1 mentions) Ly6c fitc, by BioLegend (1 mentions) Live dead fixable blue dead cell stain, by Thermo Fisher Scientific (1 mentions) Dylight 800, by Thermo Fisher Scientific (1 mentions) Cd11b buv395, by BD (1 mentions)

Spelling variants of this product

Anti-CCR2 APC (3 citations) APC-conjugated CCR2 (3 citations) APC conjugated anti-CCR2 (2 citations) Mouse CCR2 APC-conjugated antibody (2 citations)

7 protocols using ccr2 apc

Multicolor Flow Cytometry Analysis

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Cells were suspended in 3% FBS 2mM EDTA in PBS and staining was performed in the presence of 2% NRS, 2% Fc block (BD), and fixable viability dye eFluor 450 (eBioscience). Cells were stained with the following antibodies from BD: CD3-APC (145-2C11), CD19- APC (1D3), CD11c-V450 or PE-Cy7 (HL3), NK1.1-APC (PK136), Ly6G-V450 (1A8), Ly6C-PerCP-Cy5.5 (AL-21), CD11b-PE (M1/70), B220-APC (RA3-6B2), CD45-APC (30-F11), SiglecF-BV421 (E50-2440), CD8-PerCP-Cy5.5 (53-6.7). And the following antibodies from eBioscience: CD103-APC (2E7), EpCAM-PE-Cy7 (G8.8), CD31-PerCP-eFluor 710 (390), MHC Class II (I-A/I-E) eFluor 450 (M5/114.15.2), FasL-APC (MFL3), CD4-APC (GK1.5), and from R&D: CCR2-APC (475301). Alexa Fluor 647 protein labeling kit (Thermo) was used to label the influenza M2 (E10) (Bourmakina and García-Sastre, 2005 (link)) antibody. Cells were fixed with 2% formaldehyde after staining and analyzed on an LSRII after gating for FSC/SSC, singlets, and live cells. Cells were quantitated by flow cytometry with AccuCount Particles (Spherotech). Sorting was performed similarly on FACSAria.

Uccellini M.B, & García-Sastre A. (2018). ISRE-Reporter Mouse Reveals High Basal and Induced Type I IFN Responses in Inflammatory Monocytes. Cell reports, 25(10), 2784-2796.e3.

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Multiparameter Flow Cytometry Immune Profiling

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Single-cell-suspensions of splenocytes and liver leukocytes were stained for cell surface markers using specific fluorophore-conjugated antibodies optimized for flow cytometry. Reagents and antibodies used in this study were LIVE/DEAD Fixable Blue Dead Cell Stain (ThermoFisher), CD11b-BUV395 (Clone: M1/70; BD Biosciences), Siglec-F-BUV615 (Clone: E50-2440; BD Biosciences), NK1.1-BUV661 (Clone: PK136; BD Biosciences), B220-BUV737 (Clone: RA3-6B2; BD Biosciences), CD8α-BUV805 (Clone: 53–6.7; BD Biosciences), MHCII(I-A/I-E)-BV510 (Clone: M5/114.15.2; BioLegend), CD4-BV570 (Clone: RM4-5; BioLegend), SCA-1-BV711 (Clone: D7; BioLegend), CD11c-BV785 (Clone: N418; BioLegend), Ly6C-FITC (Clone: HK1.4; BioLegend), Siglec-H-PerCP/Cy5.5 (Clone: 551; BioLegend), F4/80-PE (Clone: BM8; BioLegend), CD3ε-PE/Cy5 (Clone: 145-2C11; ThermoFisher), CD80-PE/Cy7 (Clone: 16-10A1; BioLegend), CD115-AF594 (Clone: AFS98; BioLegend), CCR2-APC (Clone: 475301; R&D Systems), CD45-AF700 (Clone: 30-F11; BioLegend), CD48-APC/Cy7 (Clone: HM48-1; BioLegend), Ly6G-Biotin (Clone: 1A8; BioLegend; with secondary antibody streptavidin-DyLight 800; ThermoFisher). Stained and fixed (4% PFA; 10 min in the dark) cells were analyzed with a Becton Dickson custom 10-laser ‘LSR-II’ flow cytometer and FlowJo software (v.10.4.1).

Jung S.R., Ashhurst T.M., West P.K., Viengkhou B., King N.J., Campbell I.L, & Hofer M.J. (2020). Contribution of STAT1 to innate and adaptive immunity during type I interferon-mediated lethal virus infection. PLoS Pathogens, 16(4), e1008525.

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Flow Cytometry Antibody Panel

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We purchased directly conjugated monoclonal antibodies for flow cytometry from Invitrogen (Waltham, MA, USA): CD4-PE, Clone GK1.5; CD11b-ef450, Clone M1/70; CD44-FITC, Clone IM7; CD45AF700, Clone 30-F11; CD206-APC, Clone MR6F3; FoxP3-ef450, Clone FJK-16S, IFN-γ-PE, Clone XMG1.2; GzmB-APC, Clone GB11; iNOS-PE, Clone CXNFT; live dead fixable dead cell stain kit, Catalogue No. L34959. BD Pharmingen (San Jose, CA, USA): CD3-AF700, Clone 17A2; CD4-Pacblue and perCPcy5.5, Clone RM4-5; CD8-FITC, perCPcy5.5, and PECY7, Clone 53-6.7; CD62L-PECY7, Clone MEL-14; H2Kd-PE, Clone 17A2; Ly6G-PE and percpcy5.5, Clone 1A8; Ly6C-APCCY7, Clone- AL-21; TNF-α PECY7, Clone MP6-XT22. Biolegend (San Diego, CA, USA): CD3-PECY7 Clone 17A2; B220-percpcy5.5, Clone RA3-6B2. R&D Systems: CCR2-APC, Clone 475301; CCL2-APC, Clone 123616. eBioscience (San Diego, CA, USA): H2Kd/Dd-ef450 Clone 34-1-25; CD4-APC, Clone GK1.5.

Mittal P., Wang L., Akimova T., Leach C.A., Clemente J.C., Sender M.R., Chen Y., Turunen B.J, & Hancock W.W. (2020). The CCR2/MCP-1 Chemokine Pathway and Lung Adenocarcinoma. Cancers, 12(12), 3723.

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Multicolor Flow Cytometry Analysis

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Uccellini M.B, & García-Sastre A. (2018). ISRE-Reporter Mouse Reveals High Basal and Induced Type I IFN Responses in Inflammatory Monocytes. Cell reports, 25(10), 2784-2796.e3.

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Multiparameter Flow Cytometry Analysis

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The erythrocyte-depleted single cell suspension from BM, inguinal lymph nodes and tumor (as described in Cell Sorting) were incubated with anti-FcγR antibody, followed by incubation with fluorescently labeled antibodies. Ly6G-PE, Ly6C-PerCP/Cy5.5, CD11b-FITC, CD4-PE/Cy7, and MHCII- PE/Cy7 were purchased from Biolegend. Gr-1-PE, Foxp3-FITC, Granzyme B-PE, MHCII-PE, CD11b-PerCP/Cy5.5, LAMP2-eF660, and CD8-AF647 were purchased from eBioscience. PD-1-APC, CCR5-APC and CCR3-PE were purchased from Myltenyi Biotec. Purified anti-CCL5 (Peprotech), secondary APC-conjugated goat anti-rabbit (Columbia Bioscience), iNOS-FITC (BD Bioscience) and CCR2-APC (R&D), CCR1 (R&D) were also acquired from different commercial sources. Data acquisition was performed using FACScan (Becton Dickinson) and analyzed via FlowJo (Tree Star, Inc).

Ban Y., Mai J., Li X., Mitchell-Flack M., Zhang T., Zhang L., Chouchane L., Ferrari M., Shen H, & Ma X. (2017). Targeting autocrine CCL5-CCR5 axis reprograms immunosuppressive myeloid cells and reinvigorates antitumor immunity. Cancer research, 77(11), 2857-2868.

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Isolation and Analysis of Human Monocyte Subsets

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Whole blood was layered onto Ficoll-Paque PLUS (GE Healthcare, Uppsala, Sweden) for density gradient centrifugation and PBMC collection. Cells were immediately stained for flow cytometry, using antibodies to human CD14 (BD, Franklin Lakes, NJ; clone M5E2), CD16 (BD; clone 3G8), and CCR2-APC (R&D Systems, Minneapolis, MN; clone 48607) or isotype matched control antibodies, as described previously.^{12 (link)} Monocyte subsets were acquired with the BD FACSCantoII flow cytometer, and analysis performed using FlowJo software (v. 10.0.8, TreeStar, Ashland, OR). For this analysis, results for CD14+ and CD14+CD16+ expressing monocytes were reported as the percentage of total monocytes; for CCR2+CD14+ and CCR2+CD14+CD16+, as a percentage of CD14+ and CD14+CD16+ monocytes, respectively.

Morgello S., Buyukturkoglu K., Murray J., Veenstra M., Berman J.W., Byrd D, & Inglese M. (2021). MR spectroscopy and diffusion imaging in people with HIV: relationships to clinical and immunologic findings. Journal of neuroimaging : official journal of the American Society of Neuroimaging, 32(1), 158-170.

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Immune Cell Profiling of Murine Lungs

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Lungs were digested for 40 min in 1 mg/ml collagenase type 4 (Worthington) 5% FBS in DMEM. Cells were then filtered through a 0.2 μm cell strainer and RBCs were lysed. Cells were suspended in 3% FBS 2 mM EDTA in PBS and staining was performed in the presence of 2% NRS, 2% Fc block (BD), and fixable viability dye eFluor 450 (ebiosciences). Cells were stained with the following antibodies from BD: Ly6C-PerCP-Cy5.5 (AL-21), CD11b-PE (M1/70), Ly6G-V450 (1A8), CD11c-V450 or PE-Cy7 (HL3), CD45.1-FITC (A20), CD45.2-PE-CF594 (104), the following antibodies from eBiosciences: MHC Class II (I-A/I-E) e450 (M5/114.15.2), FasL-APC (MFL3), and from R&D: CCR2-APC (475301). Influenza M2 (E10) (85) was conjugated to Alexa 647. Cells were fixed with 2% formaldehyde after staining and analyzed on an LSRII after gating for FSC/SSC, singlets, and live cells.

Fourati S., Jimenez‐Morales D., Hultquist J.F., Chang M.W., Benner C., Krogan N.J., Pache L., Chanda S.K., Sékaly R., García‐Sastre A., & Uccellini M.B. (2022). Type I IFN promotes pathogenic inflammatory monocyte maturation during H5N1 infection.

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