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Anti cxcr5 2g8

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Anti-CXCR5 (2G8) is a monoclonal antibody that specifically binds to the CXCR5 chemokine receptor. CXCR5 is a G protein-coupled receptor that plays a role in the homing and movement of B cells.

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Lab products found in correlation

Anti biotin and anti allophycocyanin cocktails and magnetic beads, by STEMCELL (2 mentions) Easysep system, by STEMCELL (2 mentions) 220 ra3 6b2, by BD (2 mentions) Fjk 16s, by BD (2 mentions) Foxp3 staining buffer set, by Thermo Fisher Scientific (2 mentions) Facsdiva software, by BD (2 mentions) Anti cd25 pc61, by Thermo Fisher Scientific (2 mentions) Anti pd1 j43, by Thermo Fisher Scientific (2 mentions) Anti cd4 rm4 5, by BioLegend (2 mentions) Anti foxp3 fjk 16s, by Thermo Fisher Scientific (2 mentions) Anti cd38, by Thermo Fisher Scientific (2 mentions) Fixable viability dye, by Thermo Fisher Scientific (2 mentions) Anti bcl6 k112 91, by BD (2 mentions) Foxp3 transcription factor staining buffer set, by Thermo Fisher Scientific (2 mentions) Anti gl7 gl 7, by Thermo Fisher Scientific (2 mentions) Facsaria, by BD (2 mentions) Facsverse, by BD (2 mentions) Anti ctla 4 uc10 4b9, by Thermo Fisher Scientific (1 mentions) Anti gitr dta 1, by Thermo Fisher Scientific (1 mentions) Anti helios 22f6, by BioLegend (1 mentions)

Spelling variants of this product

Anti-CXCR5 (9 citations) CXCR5 (2G8, (6 citations) Biotin-conjugated anti-CXCR5 (2G8) and FITC-conjugated anti-TCRβ (H57–597) mAbs (2 citations) Anti-mouse CXCR5-biotin (2G8) (2 citations)

13 protocols using anti cxcr5 2g8

1

Enrichment and Analysis of Murine Immune Cells

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Single cell suspensions were prepared from pooled murine spleens and lymph nodes (axillary, brachial, inguinal, cervical, mesenteric, pancreatic, and para-aortic) or thymuses by mechanical disruption. Cells were stained for 1 hour at room temperature with allophycocyanin-conjugated tetramers. In some experiments cells were also stained with anti-CXCR5 (2G8, BD) antibody or phycoerythrin-conjugated tetramers. Tetramer-binding cells were enriched on magnetized columns as described previously58 (link). For thymic epithelial cell and dendritic cell analysis, thymuses were harvested and enzymatically digested as previously described59 (link). Single cell suspensions were stained with a biotinylated antibody to CD11c (HL3, BD) and an allophycocyanin-labeled antibody to EpCAM (G8.8, Biolegend). Cells were incubated with anti-biotin and anti-allophycocyanin cocktails and magnetic beads (StemCell Technologies). Cells were enriched using the EasySep system according to the manufacturer’s instructions (StemCell Technologies).
Malhotra D., Linehan J.L., Dileepan T., Lee Y.J., Purtha W.E., Lu J.V., Nelson R.W., Fife B.T., Orr H.T., Anderson M.S., Hogquist K.A, & Jenkins M.K. (2016). Polyclonal CD4+ T cell tolerance is established by distinct mechanisms, according to self-peptide expression patterns. Nature immunology, 17(2), 187-195.
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2

Multiparametric Flow Cytometry Immunophenotyping

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Fluorescence dye labeled Abs specific for CD4 (L3T4), B220 (RA3-6B2), CD44 (IM7), Fas (15A7), IgM (II/41), T- and B-cell activation antigen (GL7), ICOS (C398.4A), PD-1 (J43), IFNγ (XMG1.2), IL-4 (11B11), IL-17A (eBio17B7), IL-21 (FFA21), Bcl-6 (K112-91) and FoxP3 (FJK-16s) were purchased from BD, eBioscience and Biolegend. Analysis of CXCR5 expression was performed using a biotinylated anti-CXCR5 (2G8, BD) antibody followed by incubation with APC- or APC.Cy7-labelled streptavidin, as described 25 (link). Intracellular staining for Bcl-6, FoxP3 and cytokines was performed using the FoxP3 staining buffer set (eBioscience). Intracellular staining of phospho-S473-AKT (M89-61), pSTAT1 (14/P-STAT1), pSTAT3 (4/P-STAT3) was conducted according to manufacturer’s instructions (BD Bioscience). Cells were acquired on a FACSCantoII using FACSDIva software (BD Biosciences) and analyzed with FlowJo software (Tristar).
Leavenworth J.W., Verbinnen B., Yin J., Huang H, & Cantor H. (2014). A p85α–osteopontin axis couples the ICOS receptor to sustained Bcl-6 expression by follicular helper and regulatory T cells. Nature immunology, 16(1), 96-106.
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3

Multiparameter flow cytometry analysis

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The following monoclonal antibodies were used for multiparameter flow cytometry: anti-CD4 (RM4-5, Biolegend), anti-CD25 (PC61.5, eBioscience), anti-CCR6 (29-2L17, Biolegend), anti-CD103 (2E7, Biolegend), anti-Ly5.1 (A20, Biolegend), anti-Ly5.2 (104, Biolegend), anti-CTLA4 (UC10-4B9, eBioscience), anti-Ki67 (solA15, eBioscience), anti-Bcl-2 (BCL10C4, Biolegend), anti-ICOS (7E.17G, eBioscience), anti-GITR (DTA-1, eBioscience), anti-Helios (22F6, Biolegend), anti-Neuropilin (FAB566A, R&D systems), anti-PD1 (J43, eBioscience), anti-CXCR5 (2G8, BD Biosciences). The Foxp3 staining kit was used for staining for CTLA-4, Ki67, Helios, Bcl-2 and Bcl6. Stained cells were analyzed on FACS Canto (Becton Dickinson) and were sorted on FACS Aria (Becton Dickinson). Data were analyzed with Flowjo software.
Chandrasekaran U., Yi W., Gupta S., Weng C.H., Giannopoulou E., Chinenov Y., Jessberger R., Weaver C.T., Bhagat G, & Pernis A.B. (2016). Regulation of effector Tregs in murine lupus. Arthritis & rheumatology (Hoboken, N.J.), 68(6), 1454-1466.
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4

Comprehensive Immune Cell Profiling

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The following fluorophore-conjugated antibodies were used for flow cytometry: anti-CD8 (53–6.7), anti-Thy1.1 (OX-7), anti-Vα2 (B20.1), anti-CD45 (30-F11), anti-PD-1 (29F.1A12), anti-CXCR3 (CXCR3–173), anti-IFNγ (XMG1.2), anti-IL-2 (JES6–5H4), anti-TNFα (MP6-XT22), anti-CD28 (37.51), anti-Tbet (4B10) (Biolegend); anti-CD27 (LG.7F9), anti-LAG3 (C9B7W), anti-granzyme B (Ngzb), anti-Eomes (Dan11mag) (Thermo Fisher); anti-pCD3ζ (pY142) (K25–407.69), anti-CXCR5 (2G8) (BD Biosciences).
Snook J.P., Soedel A.J., Ekiz H.A., O’Connell R.M, & Williams M.A. (2020). Inhibition of SHP-1 expands the repertoire of antitumor T cells available to respond to immune checkpoint blockade.. Cancer immunology research, 8(4), 506-517.
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5

Multicolor Flow Cytometry Staining

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Anti-CXCR5 (2G8) and GL7 (GL7) Abs were from BD Biosciences. Fixable viability dye, anti-CD38 and anti-Foxp3 (FJK-16s) Abs were from eBioscience. Anti-CD4 (GK1.5), anti-B220 (RA3-6B2), anti-IgG1 (RMG1-1), anti-PD-1 (29F.1A12) were from Biolegend.
Xie M.M., Koh B.H., Hollister K., Wu H., Sun J., Kaplan M.H, & Dent A.L. (2017). Bcl6 Promotes Follicular Helper T Cell Differentiation and PD-1 Expression in a Blimp1-independent Manner. European journal of immunology, 47(7), 1136-1141.
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6

Multiparametric Flow Cytometry Immunophenotyping

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Fluorescence dye labeled Abs specific for CD4 (L3T4), B220 (RA3-6B2), CD44 (IM7), Fas (15A7), IgM (II/41), T- and B-cell activation antigen (GL7), ICOS (C398.4A), PD-1 (J43), IFNγ (XMG1.2), IL-4 (11B11), IL-17A (eBio17B7), IL-21 (FFA21), Bcl-6 (K112-91) and FoxP3 (FJK-16s) were purchased from BD, eBioscience and Biolegend. Analysis of CXCR5 expression was performed using a biotinylated anti-CXCR5 (2G8, BD) antibody followed by incubation with APC- or APC.Cy7-labelled streptavidin, as described 25 (link). Intracellular staining for Bcl-6, FoxP3 and cytokines was performed using the FoxP3 staining buffer set (eBioscience). Intracellular staining of phospho-S473-AKT (M89-61), pSTAT1 (14/P-STAT1), pSTAT3 (4/P-STAT3) was conducted according to manufacturer’s instructions (BD Bioscience). Cells were acquired on a FACSCantoII using FACSDIva software (BD Biosciences) and analyzed with FlowJo software (Tristar).
Leavenworth J.W., Verbinnen B., Yin J., Huang H, & Cantor H. (2014). A p85α–osteopontin axis couples the ICOS receptor to sustained Bcl-6 expression by follicular helper and regulatory T cells. Nature immunology, 16(1), 96-106.
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7

Multiparameter Analysis of Immune Cells

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Single-cell suspensions were prepared from the spleen, lymph nodes (LNs), or peripheral blood lymphocytes (PBLs), and surface or intracellularly stained as described (19 (link)). The fluorochrome-conjugated antibodies were as follows: anti-CD8 (53-6.7), anti-CD4 (RM4-5), anti-CD44 (IM7), anti-CD62L (MEL-14), anti-PD-1 (J43), anti-CD45.1 (A20), anti-CD45.2 (104), anti-ICOS (C398.4A), anti-IFN-γ (XMG1.2), anti-TNFα (MP6-XT22), anti-Eomes (Dan11mag), anti-T-bet (eBio4B10), anti-IL-2 (JES6-5H4), anti-IL-7Rα (eBio17B7) and anti-KLRG1 (2F1) from eBiosciences; anti-Bcl6 (K112-91) from BD Biosciences; anti-human granzyme B (FGB12) and corresponding isotype control from Invitrogen/Life Technologies, anti-SLAM (TC15-12F12.2) from BioLegend. For detection of CXCR5, three-step staining protocol was used with unconjugated anti-CXCR5 (2G8; BD Biosciences) (24 (link)). For detection of Bcl6, surface-stained cells were fixed and permeabilized with the Foxp3/Transcription Factor Staining Buffer Set (eBiosciences), followed by incubation with a fluorochrome-conjugated antibody or isotype control. Peptide-stimulated cytokine production and detection by intracellular staining were as described (25 (link)). Data were collected on a FACSVerse (BD Biosciences) and were analyzed with FlowJo software (TreeStar).
Gullicksrud J.A., Li F., Xing S., Zeng Z., Peng W., Badovinac V.P., Harty J.T, & Xue H.H. (2017). Differential requirements for Tcf1 long isoforms in CD8+ and CD4+ T cell responses to acute viral infection. Journal of immunology (Baltimore, Md. : 1950), 199(3), 911-919.
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8

Enrichment and Analysis of Murine Immune Cells

Cited 1 time
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Single cell suspensions were prepared from pooled murine spleens and lymph nodes (axillary, brachial, inguinal, cervical, mesenteric, pancreatic, and para-aortic) or thymuses by mechanical disruption. Cells were stained for 1 hour at room temperature with allophycocyanin-conjugated tetramers. In some experiments cells were also stained with anti-CXCR5 (2G8, BD) antibody or phycoerythrin-conjugated tetramers. Tetramer-binding cells were enriched on magnetized columns as described previously58 (link). For thymic epithelial cell and dendritic cell analysis, thymuses were harvested and enzymatically digested as previously described59 (link). Single cell suspensions were stained with a biotinylated antibody to CD11c (HL3, BD) and an allophycocyanin-labeled antibody to EpCAM (G8.8, Biolegend). Cells were incubated with anti-biotin and anti-allophycocyanin cocktails and magnetic beads (StemCell Technologies). Cells were enriched using the EasySep system according to the manufacturer’s instructions (StemCell Technologies).
Malhotra D., Linehan J.L., Dileepan T., Lee Y.J., Purtha W.E., Lu J.V., Nelson R.W., Fife B.T., Orr H.T., Anderson M.S., Hogquist K.A, & Jenkins M.K. (2016). Polyclonal CD4+ T cell tolerance is established by distinct mechanisms, according to self-peptide expression patterns. Nature immunology, 17(2), 187-195.
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9

Multi-color Flow Cytometry Analysis

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Fluorochrome-labeled monoclonal antibodies against CD4 (GK1.5), CD8a (53-6.7), TCRβ (H57-597), CD44 (IM7), CD25 (PC61), B220 (RA3-6B2), CXCR5 (2G8), PD1 (29F.1A12, J43), ICOS (C398.4A), Fas (JO2), GL7 (GL7), CD3e (17A2), CD45.1 (A20), CD45.2 (104), CD19 (ID3), IgD (11-26c.2a), CD69 (H1.2F3), γδ-TCR (GL3), Vα2 (B20.1), Vβ5 (MR9-4), CXCR5 (L138D7), BTLA (HMBT-6B2), and OX40 (OX86) were purchased from e-Bioscience, BD, or BioLegend. We used either fluorophore-conjugated CXCR5 antibody (L138D7) or purified CXCR5 (2G8) antibody. For CXCR5 staining using purified antibody, cells were first incubated with 5 µg/ml of purified anti-CXCR5 (2G8; BD), followed by biotinylated AffiniPure anti–rat IgG (Jackson ImmunoResearch Laboratories, Inc.) and APC-streptavidin (Invitrogen). After addition of 100 µg/ml of purified rat IgG2a isotype, cells were incubated with other extracellular antibodies. Bcl6 (clone K12-91; BD), Batf (clone MBM7C7; eBioscience), Foxp3 (clone FJK-16s; eBioscience), Tbet (clone 4B10; BioLegend), Ki-67 (clone SolA15; eBioscience), and Gata3 (TWAJ; eBioscience) were stained after fixation and permeabilization with the Foxp3 staining buffer set from eBioscience. Samples were analyzed on an LSRII (BD) or sorted on a FACSAria (BD) or MoFlo (Dako). Data were analyzed using FlowJo software (Tree Star).
Mathew R., Mao A.P., Chiang A.H., Bertozzi-Villa C., Bunker J.J., Scanlon S.T., McDonald B.D., Constantinides M.G., Hollister K., Singer J.D., Dent A.L., Dinner A.R, & Bendelac A. (2014). A negative feedback loop mediated by the Bcl6–cullin 3 complex limits Tfh cell differentiation. The Journal of Experimental Medicine, 211(6), 1137-1151.
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10

Comprehensive Immune Cell Staining

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The following staining reagents were used: anti-CD3 (145-2C11), anti-CD4 (H129.19), anti-CD62L (MEL-14), anti-CD44 (IM7), anti-CD45R (RA3-6B2), anti-PD-1 (29F.1A12), anti-PD-L1 (10F.9G2), anti-ICOSL (HK5.3), anti-CD40 (HM40-3), anti-CD40L (MR1), anti-CXCR4 (L276F12), and anti-IL-4 (11b11) were from Biolegend (San Diego, CA, USA); anti-CD19 (eBio1D3), anti-ICOS (7E.17G9), anti-IL-21 (mhalx21), anti-CD69 (H1.2F3), and anti-GL7 (GL-7) were from eBiosciences (San Diego, CA, USA); anti-Bcl-6 (K112-91), anti-CXCR5 (2G8), and anti-Fas (Jo2) were from BD Biosciences (Franklin Lakes, NJ, USA). PNA (B-1075; Vector Laboratories, Burlington, ON, Canada) was used for staining germinal center B cells and MOMA-1 (Abcam, Cambridge, UK) for staining MZ macrophages.
Daugan M., Murira A., Mindt B.C., Germain A., Tarrab E., Lapierre P., Fritz J.H, & Lamarre A. (2016). Type I Interferon Impairs Specific Antibody Responses Early during Establishment of LCMV Infection. Frontiers in Immunology, 7, 564.
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