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|67761S||100 µl (50 tests)||$329.00.0|
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TIM-3 (D5D5R™) XP® Rabbit mAb (PE Conjugate) #67761
Flow cytometric analysis of Jurkat cells (blue) and primary CD4+ T cells (green) using TIM-3 (D5D5R™) XP® Rabbit mAb (PE Conjugate). Controls were run on Jurkat cells (orange) and primary CD4+ T cells (red) using concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control (PE Conjugate) #5742. CD4+ T cells were purified from human blood and stimulated for 9 days using beads coated with CD3 and CD28 antibodies in the presence of Human Interleukin-2 (hIL-2) #8907 (6.7 ng/ml).Learn more about how we get our images
Gallery: TIM-3 (D5D5R™) XP® Rabbit mAb (PE Conjugate) #67761
Flow Cytometry Protocol for TIM-3 (D5D5R) XP® Rabbit mAb (PE Conjugate) #67761
A. Solutions and Reagents
- 1X Phosphate Buffered Saline (PBS): Dissolve 8 g NaCl, 0.2 g KCl, 1.44 g Na2HPO4, and 0.24 g KH2PO4 in 800 ml distilled water (dH2O). Adjust the pH to 7.4 with HCl and the volume to 1 liter with dH2O. Store at room temperature.
- Incubation Buffer: Dissolve 0.5 g bovine serum albumin (BSA) in 100 ml 1X PBS. Store at 4°C.
NOTE: Account for isotype matched controls for monoclonal antibodies or species matched IgG for polyclonal antibodies. Count cells using a hemacytometer or alternative method.
- Collect cells by centrifugation and aspirate supernatant.
- Aliquot 0.5–1 x 106 cells into each assay tube (by volume).
- Add 2–3 ml Incubation Buffer to each tube and rinse by centrifugation. Repeat.
- Resuspend cells in 100 µl Incubation Buffer per assay tube.
- Block in Incubation Buffer for 10 minutes on ice.
- Add the conjugated antibody at the appropriate dilution to the assay tubes (see individual antibody data sheet for the dilution recommendations).
- Incubate for 1 hour on ice.
- Rinse as before in Incubation Buffer by centrifugation.
- Resuspend cells in 0.5 ml PBS and analyze on flow cytometer.
revised April 2016
TIM-3 (D5D5R™) XP® Rabbit mAb (PE Conjugate) recognizes endogenous levels of total TIM-3 protein.Species Reactivity: Human
Monoclonal antibody is produced by immunizing animals with recombinant protein specific to the extracellular domain of human TIM-3 protein.
This Cell Signaling Technology antibody is conjugated to phycoerythrin (PE) and tested in-house for direct flow cytometry analysis in human cells. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated TIM-3 (D5D5R™) XP® Rabbit mAb #45208.
T cell Ig- and mucin-domain-containing molecules (TIMs) are a family of transmembrane proteins expressed by various immune cells. TIM-3 is an inhibitory molecule that is induced following T cell activation (1-3 ). TIM-3 is expressed by exhausted T cells in the settings of chronic infection and cancer (4,5), and tumor-infiltrating T cells that coexpress PD-1 and TIM-3 exhibit the most severe exhausted phenotype (5). Tumor-infiltrating dendritic cells (DCs) also express TIM-3. TIM-3 expression on DCs was found to suppress innate immunity by reducing the immunogenicity of nucleic acids released by dying tumor cells (6). Research studies show that heterodimerization of TIM-3 with CEACAM-1 is critical for the inhibitory function of TIM-3, and co-blockade of TIM-3 and CEACAM-1 enhanced antitumor responses in a mouse model of colorectal cancer (7). In addition, blockade of TIM-3 in mouse models of autoimmunity enhanced the severity of disease (1). Finally, binding of Galectin-9 to TIM-3 expressed by Th1 cells induces T cell death (8).
For Research Use Only. Not For Use In Diagnostic Procedures. Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. XP is a registered trademark of Cell Signaling Technology, Inc.