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Flow cytometric analysis of CCR7 (CD197) on Human peripheral blood leukocytes and lymphocytes. Whole blood was stained with BD Horizon™ BUV395 Mouse Anti-Human CD4 (clone RPA-T4, Cat. No. 564724), APC Mouse Anti-Human CD8 (clone RPA-T8, Cat. No. 561952, 561421, 555369, or 561953), FITC Mouse Anti-Human CD45RA (clone HI100, Cat. No. 561882 or 555488), and either BD Horizon™ BV605 Mouse IgG1, κ Isotype Control (Cat. No. 562652) or BD Horizon™ BV605 Mouse Anti-Human CCR7 (CD197) (Cat. No. 566754 or 566755). The erythrocytes were lysed with BD PharmLyse™ Lysing Buffer (Cat. No. 555899). Flow cytometry and data analysis were performed using a BD FACS LSRFortessa™ X--20 Flow Cytometer and FlowJo™ Software. Plots 1 and 2: The contour plots showing Ig isotype control or CCR7 (CD197) staining versus side light-scatter were derived from gated events with the forward and side light-scatter characteristics of viable leukocytes. Plot 3: The contour plot showing CCR7 (CD197) staining versus CD45RA was derived from CD4-positive T cell gated events with the forward and side light-scatter characteristics of viable lymphocytes. Please note that we have observed considerable donor-to-donor variation in the proportion of CCR7 (CD197)-negative CD4-positive lymphocytes. Plot 4: The contour plot showing CCR7 (CD197) staining versus CD45RA was derived from CD8-positive T cell gated events with the forward and side light-scatter characteristics of viable lymphocytes.
BD Horizon™ BV605 Mouse Anti-Human CCR7 (CD197)
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Recommended Assay Procedures
BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
For optimal and reproducible results, BD Horizon Brilliant™ Stain Buffer should be used anytime BD Horizon Brilliant dyes are used in a multicolor flow cytometry panel. Fluorescent dye interactions may cause staining artifacts which may affect data interpretation. The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. When BD Horizon Brilliant Stain Buffer is used in the multicolor panel, it should also be used in the corresponding compensation controls for all dyes to achieve the most accurate compensation. For the most accurate compensation, compensation controls created with either cells or beads should be exposed to BD Horizon Brilliant Stain Buffer for the same length of time as the corresponding multicolor panel. More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).
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Companion Products
The 2-L1-A monoclonal antibody specifically binds to the human CC chemokine receptor CCR7, also known as CD197, on the cell surface. CCR7 (previously known as BLR2, EBI1 and CMKBR7) is a seven-transmembrane, G-protein-coupled receptor specific for two CC chemokines: CCL19 (also known as MIP-3β, Exodus-3, and ELC) and CCL21 (also known as 6Ckine, Exodus-2 SLC, TCA4, and SCYA21). CCR7 mRNA is expressed mainly in lymphoid tissues including the spleen, lymph nodes and tonsil, in bone marrow, and on peripheral T and B lymphocytes, cord blood CD34-positive cells, and mature dendritic cells. In response to its cognate chemokines, CCR7 (CD197) mediates homing of leucocytes to secondary lymphoid tissues. Differential CCR7 (CD197) expression can be used to distinguish naive, central memory, and effector memory T cell subsets. The human CCR7 gene, unlike other CC chemokine receptor genes, has been mapped to chromosome 17 (region 17q12). Because the extracellular region of CCR2 (CD192) has significant sequence homology with CCR7 (CD197), BD Biosciences has confirmed that mAb 2-L1-A does not cross-react with CCR2 on the surface of transfected cells.
Development References (8)
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