Flow cytometric analysis of live human peripheral blood mononuclear cells, untreated (blue) or treated with TPA (12-O-Tetradecanoylphorbol-13-Acetate) #4174 (0.081 uM, 5 hr) and Ionomycin, Calcium Salt #9995 (1.34 uM, 5 hr; green), using CD69 (FN50) Mouse mAb (PerCP-Cy5.5® Conjugate) (solid lines) or concentration-matched Mouse (MOPC-21) mAb IgG1 Isotype Control (PerCP-Cy5.5® Conjugate) #24589 (dashed lines).
|Source/Isotype||Mouse IgG1 kappa|
This Cell Signaling Technology antibody is conjugated to PerCP-Cy5.5® and tested in-house for direct flow cytometric analysis in human cells.
Supplied in 10 mM NaH2PO4, 150 mM NaCl, 0.09% NaN3, 0.1% gelatin, pH 7.2. This product is stable for 6 months when stored at 4ºC. Do not aliquot the antibody. Protect from light. Do not freeze.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: When including fluorescent cellular dyes in your experiment (including viability dyes, DNA dyes, etc.), please refer to the dye product page for the recommended protocol. Visit www.cellsignal.com for a full listing of cellular dyes validated for use in flow cytometry.
NOTE: Count cells using a hemocytometer or alternative method.
NOTE: If using whole blood, lyse red blood cells and wash by centrifugation prior to Immunostaining.
NOTE: Optimal centrifugation conditions will vary depending upon cell type and reagent volume. Generally, 150-300g for 1-5 minutes will be sufficient to pellet the cells.
posted June 2017
revised June 2020
Protocol Id: 1504
CD69 (FN50) Mouse mAb (PerCP-Cy5.5® Conjugate) recognizes endogenous levels of total CD69 protein. This antibody detects an epitope within the extracellular domain.
This monoclonal antibody was purified from tissue culture supernatant via affinity chromatography. The purified antibody was conjugated under optimal conditions, with unreacted dye removed from the preparation.
CD69, also known as Leu-23, is a type II transmembrane glycoprotein that is expressed on the surface of T cells, B cells, and NK cells (1,2). This phosphorylated disulfide-linked 28 to 32-kDa homodimer is constitutively expressed on a subset of thymocytes and platelets. It also acts as an activation antigen of lymphocytes, NK cells, neutrophils, and eosinophils (1-6). Studies have shown that stimulation of the T cell receptor (TCR) increases the expression of CD69 on the cell surface. The ability to detect the level of CD69 expression after TCR activation makes CD69 an ideal indicator of T cell activation (1).
The FN50 antibody is widely used as a marker for T cell activation (7).
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