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Flow cytometric analysis of CD185 (CXCR5) expression on mouse splenocytes. Splenocytes were stained with FITC Rat Anti-Mouse CD45R/B220 (Cat. No. 553088) and Purified Rat Anti-Mouse CD185 (CXCR5) (Cat. No. 551961), followed by Biotin Mouse Anti-Rat IgG2a (Cat. No. 553894) and PE Streptavidin (Cat. No. 554061). Two-color dot plot depicting CD185 (CXCR5) and CD45R/B220 expression were derived from gated events with the side and forward light-scattering characteristics of viable lymphocytes. The quadrant markers for the bivariate dot plots were set based on the Purified Rat IgG2a, κ Isotype Control (Cat. No. 553927) staining.
Flow cytometric analysis of CD185 (CXCR5) expression on mouse splenocytes. Splenocytes were stained with FITC Rat Anti-Mouse CD45R/B220 (Cat. No. 553088) and Purified Rat Anti-Mouse CD185 (CXCR5) (Cat. No. 551961), followed by Biotin Mouse Anti-Rat IgG2a (Cat. No. 553894) and PE Streptavidin (Cat. No. 554061). Two-color dot plot depicting CD185 (CXCR5) and CD45R/B220 expression were derived from gated events with the side and forward light-scattering characteristics of viable lymphocytes. The quadrant markers for the bivariate dot plots were set based on the Purified Rat IgG2a, κ Isotype Control (Cat. No. 553927) staining.
Flow cytometric analysis of CD185 (CXCR5) expression on mouse splenocytes. Splenocytes were stained with FITC Rat Anti-Mouse CD45R/B220 (Cat. No. 553088) and Purified Rat Anti-Mouse CD185 (CXCR5) (Cat. No. 551961), followed by Biotin Mouse Anti-Rat IgG2a (Cat. No. 553894) and PE Streptavidin (Cat. No. 554061). Two-color dot plot depicting CD185 (CXCR5) and CD45R/B220 expression were derived from gated events with the side and forward light-scattering characteristics of viable lymphocytes. The quadrant markers for the bivariate dot plots were set based on the Purified Rat IgG2a, κ Isotype Control (Cat. No. 553927) staining.
Any use of products other than the permitted use without the express written authorization of Becton, Dickinson and Company is strictly prohibited.
A multistep staining procedure is recommended to amplify immunofluorescent signals for the flow cytometric analysis of mouse CXCR5 expression:
Step 1: Incubate 1 million cells with 0.1 - 0.5 µg of Purified Rat Anti-Mouse CXCR5 at 4°C for 15 - 20 minutes. Wash cells two times with staining medium containing sodium azide (e.g., Dulbecco's PBS or tissue culture medium [without phenol red and biotin] with 0.09% sodium azide and 2% heat-inactivated FCS or 0.2% BSA).
Step 2: Incubate the cells with Biotin Mouse Anti-Rat IgG2a (Cat. No. 553894) at 4°C for 20 minutes. Wash cells two times.
Step 3 : Incubate the cells with ≤ 0.06 µg of PE Streptavidin (Cat. No. 554061) at 4°C for 20 minutes. Wash two times. Resuspend cells in staining medium and analyze stained cells with a FACScan™ Flow Cytometer (Becton Dickinson, San Jose, CA) using appropriate specificity and compensation controls.
The 2G8 monoclonal antibody specifically binds to the mouse C-X-C Chemokine Receptor type 5, CXCR5. CXCR5 is also known as CD185, BLR1, NLR and MDR15. CXCR5 is a seven-transmembrane, G-protein-coupled receptor that is specific for the CXC chemokine, CXCL13/BLC/BCA-1. The expression of CXCR5 has been detected in spleen, lymph nodes, tonsils, brain, bone marrow, T cells, B cells, cerebrum, cerebellum, hippcampus and pituitary. In mouse spleen, CXCR5 was strictly expressed by mature B cells and a small subset of T lymphocytes. CXCR5 plays a role in directing the migration of B and T cells to B cell follicles with the spleen and certain other lymphoid tissues. The immunogen used to generate 2G8 hybridoma was a recombinant protein containing N-terminal amino acids of mouse CXCR5 (GST-NmBLR1).
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