Flow cytometric analysis of live human peripheral blood mononuclear cells using KIR2DL3 (D8L3D) Rabbit mAb compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900. Samples were co-stained with CD56 to distinguish NK cell population. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412. Total viable cells were used for analysis.
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
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.
revised June 2020
Protocol Id: 1865
KIR2DL3 (D8L3D) Rabbit mAb recognizes endogenous levels of total KIR2DL3 protein. This antibody weakly cross-reacts with KIR2DL2 proteins in over-expression cell lines.
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ala173 of human KIR2DL3 protein.
Killer cell immunoglobulin-like receptors (KIRs) are type 1 transmembrane glycoproteins expressed by natural killer cells and subsets of CD4, CD8, and γδ T cells (1-5). Analogous to the diversity of their human leucocyte antigen class I (HLA Class I) ligands, the KIR genes are polymorphic and the content of the KIR gene cluster varies among haplotypes, although several "framework" genes are found in all haplotypes (6-7). The KIR proteins are characterized by the number of extracellular immunoglobulin-superfamily domains (2D or 3D) and by whether they have a long (L) or short (S) cytoplasmic domain (8-10). KIR proteins with the long cytoplasmic domain transduce inhibitory signals upon ligand binding via an immune tyrosine-based inhibitory motif (ITIM) (10), while KIR proteins with the short cytoplasmic domain lack an ITIM and instead transduce activating signals (11,12). KIR proteins play an important role in the regulation of the immune response. Combinations of KIR and HLA class I variants influence susceptibility to autoimmunity and infectious disease, as well as outcomes of haematopoietic stem cell transplantation (12-14).
KIR2DL3, also referred to as CD158b, interacts with HLA-C alleles (HLA-Cw1, HLA-Cw3, and HLA-Cw7). Upon receptor ligand interaction, KIR2DL3 inhibits the activity of NK cells thus preventing target cell lysis (15-17).
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