Anti gl7 gl 7

Manufactured by Thermo Fisher Scientific

Sourced in United States

Anti-GL7 (GL-7) is a laboratory reagent used in flow cytometry applications. It is a monoclonal antibody that specifically binds to the GL7 antigen, which is expressed on activated B cells and germinal center B cells. The core function of Anti-GL7 (GL-7) is to facilitate the identification and analysis of these cell populations in biological samples.

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5 protocols using anti gl7 gl 7

Phenotyping Immune Cell Subsets

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Surface staining of cells was performed by suspending them in blocking solution (10% fetal calf serum in PBS) for 30 min at 4°C in the presence of the following fluorescent-labeled mAbs: anti-B220 (RA3-6B2, eBioscience), anti-CD3 (145-2C11, TONBO Bioscience), anti-CD4 (GK1.5, BioLegend), anti-CD8 (53-6.7, TONBO Bioscience), anti-CD25 (PC61.5, eBioscience), anti-CD21 (7G6, eBioscience), anti-CD23 (B3B4, eBioscience) and anti-LAP (TW7-16B4, BioLegend), anti-CXCR5 (SPRCL5, eBioscience), anti-PD1 (J43, eBioscience), anti-CD138 (281-2, BioLegend), anti-CD19 (1D3, TONBO Bioscience), anti-GL-7 (GL7, eBioscience), anti-IgM (eb121-15-F9, eBioscience), anti-IgD (11-26c, eBioscience), anti-CD40L (MR1, eBioscience), and anti-CD69 (H1.2F3, BioLegend). For intracellular cytokine staining, cells were exposed to monensin, fixed and permeabilized using the BD Cytofix/Cytoperm Plus Kit with BD GolgiStop™ (BD Biosciences) following manufacturer’s indications, and then stained with specific fluorescent-labeled mAbs: anti-IFN-γ (FITC-XMG1.2, TONBO Bioscience) and anti-IL-2 (PE-JES6-5H4, BD Pharmingen). For FoxP3 staining, the Anti-Mouse/Rat Foxp3 PE Staining Set (eBioscience) was used following manufacturer’s indications. Flow cytometry analyses were performed on a FACS Canto II equipped with CellQuest (BD Biosciences) and Flowjo 8.7 software.

Consuegra-Fernández M., Martínez-Florensa M., Aranda F., de Salort J., Armiger-Borràs N., Lozano T., Casares N., Lasarte J.J., Engel P, & Lozano F. (2017). Relevance of CD6-Mediated Interactions in the Regulation of Peripheral T-Cell Responses and Tolerance. Frontiers in Immunology, 8, 594.

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Flow Cytometry Analysis of Bone Marrow and Spleen Leukocytes

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Single-cell leukocyte suspensions from the bone marrow and spleen were used in the flow cytometry experiments. CD16/CD32 (93, BioLegend) was used to block FcRs. The following antibodies [conjugated to eFluor 450, fluorescein isothiocyanate, phycoerythrin (PE), PE-Cy5, PerCP/Cy5.5, and PE-Cy7] were used: anti-CD21 (7G6, BD Biosciences), anti-CD23 (B3B4, eBioscience), anti-CD43 (S7, BD Biosciences), anti-CD93 (AA4.1, eBioscience), anti-CD95 (15A7, eBioscience), anti-B220 (RA3-6B2, BD Biosciences), anti-GL7 (GL-7, eBioscience), anti-IgD (11-26c.2a, BD Biosciences), and anti-IgM (II/41, eBioscience). Detection of cell surface marker expression was performed using an LSRFortessa cytometer (BD Biosciences). Typically, living lymphocytes, judged by forward and side scatter parameters, were gated for analysis. Data were further analyzed using FlowJo (Tree Star).

Zhao X., Xie H., Zhao M., Ahsan A., Li X., Wang F., Yi J., Yang Z., Wu C., Raman I., Li Q.Z., Kim T.J, & Liu W. (2019). Fc receptor–like 1 intrinsically recruits c-Abl to enhance B cell activation and function. Science Advances, 5(7), eaaw0315.

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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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Cryosection Immunofluorescence Protocol

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Cryosections 7 µm in thickness from mLNs and spleen were cut and prepared as described previously (Green et al., 2011 (link)). Primary antibodies used for staining cryosections were biotinylated anti-Thy1.1 (CD90.1; HIS51; eBioscience), biotinylated anti-GL7 (GL7; eBioscience), biotinylated or unlabeled anti-CD35 (8C12; BD), and unlabeled polyclonal goat anti–mouse IgD (Cedarlane). Secondary antibodies or streptavidin fused to alkaline phosphatase or peroxidase were from Jackson ImmunoResearch Laboratories, Inc. Images were captured with an AxioObserver Z1 inverted microscope (Carl Zeiss).

Muppidi J.R., Lu E, & Cyster J.G. (2015). The G protein–coupled receptor P2RY8 and follicular dendritic cells promote germinal center confinement of B cells, whereas S1PR3 can contribute to their dissemination. The Journal of Experimental Medicine, 212(13), 2213-2222.

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Flow Cytometric Analysis of Spleen Cells

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Flow cytometric analysis was carried out on the spleens of the indicated animals. Spleens were passed through a 100μm cell strainer (Corning) and the red cells lysed. Samples were resuspended, FC receptors blocked (2.4G2, eBioscience) and surface antigens stained in HBSS (Life Technologies) alone or PBS (Life technologies) plus 0.5% (w/v) BSA (Sigma). The Foxp3 Transcription Factor Staining Buffer Set (eBioscience) was used for intracellular staining. The following stains and antibodies were used: Live/dead (Life Technologies), anti-CD4 (GK1.5, Tonbo Biosciences), anti-CD8 (53-67, Tonbo Bioscience), anti-CD11c (N418, eBioscience), anti-CD19 (6D5, eBioscience), anti-CD25 (PC61.5, eBioscience), anti-CD40L (MR1, eBioscience), anti-CD44 (IM7, eBioscience), anti-CD62L (MEL14, eBioscience), anti-CXCR5 (2G8, BD Bioscience), anti-FoxP3 (FJK-16s, eBioscience), PNA (Vector Laboratories), anti-GL7 (GL-7, eBioscience), anti-ICOS (7E.17G9, eBioscience), anti-IgD (11-26c, eBioscience), anti-IgM (eb121-15F9), anti-PD1 (J43, eBioscience), anti-DR3 (BAF2437, R&D) and anti-TL1A (Tan2-2, University of Southampton [19 (link)]). Samples were run for analysis on a BD FACSVerse and sorted on a BD FACS Aria II. Flow cytometrey data was analyzed with Flow Jo V9.

Ferdinand J.R., Richard A.C., Meylan F., Al-Shamkhani A, & Siegel R.M. (2018). Cleavage of TL1A differentially regulates its effects on innate and adaptive immune cells. Journal of immunology (Baltimore, Md. : 1950), 200(4), 1360-1369.

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