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95663
4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb (PE Conjugate)
Antibody Conjugates
Monoclonal Antibody

4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb (PE Conjugate) #95663

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Flow Cytometry Image 1 - 4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb (PE Conjugate)

Flow cytometric analysis of human peripheral blood mononuclear cells, untreated (left) or treated with anti-CD3 plus anti-CD28 (10 μg/ml, 72 hr) (right), using 4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb (PE Conjugate) (top row) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control (PE Conjugate) #5742 (bottom row) and co-stained with a CD8 antibody.

Flow Cytometry Image 2 - 4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb (PE Conjugate)

Flow cytometric analysis of live Jurkat cells (blue, negative) and HDLM-2 cells (green, positive) using 4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb (PE Conjugate) (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control (PE Conjugate) #5742 (dashed lines).

To Purchase # 95663S
Product # Size Price
95663S
100 µl  (50 tests) $ 314

Supporting Data

REACTIVITY H
SENSITIVITY Endogenous
MW (kDa)
Source/Isotype Rabbit IgG

Application Key:

  • W-Western
  • IP-Immunoprecipitation
  • IHC-Immunohistochemistry
  • ChIP-Chromatin Immunoprecipitation
  • IF-Immunofluorescence
  • F-Flow Cytometry
  • E-P-ELISA-Peptide

Species Cross-Reactivity Key:

  • H-Human
  • M-Mouse
  • R-Rat
  • Hm-Hamster
  • Mk-Monkey
  • Mi-Mink
  • C-Chicken
  • Dm-D. melanogaster
  • X-Xenopus
  • Z-Zebrafish
  • B-Bovine
  • Dg-Dog
  • Pg-Pig
  • Sc-S. cerevisiae
  • Ce-C. elegans
  • Hr-Horse
  • All-All Species Expected

Product Description

This Cell Signaling Technology antibody is conjugated to phycoerythrin (PE) and tested in-house for direct flow cytometric analysis in human cells. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated 4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb #34594.

Product Usage Information

Application Dilution
Flow Cytometry 1:50

Storage

Supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibodies. Protect from light. Do not freeze.

Protocol

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Flow Cytometry, Live Cell Protocol for Directly Conjugated Antibodies

A. Solutions and Reagents

NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.

  1. 1X Phosphate Buffered Saline (PBS): To prepare 1 L 1X PBS: add 100 ml 10X PBS (#12528) to 900 ml water, mix.
  2. Antibody Dilution Buffer: Purchase ready-to-use Flow Cytometry Antibody Dilution Buffer (#13616), or prepare a 0.5% BSA PBS buffer by dissolving 0.5 g Bovine Serum Albumin (BSA) (#9998) in 100 ml 1X PBS. Store at 4°C.

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.

B. Immunostaining

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.

  1. Aliquot desired number of cells into tubes or wells. (Generally, 5x105 to 1x106 cells per assay.)
  2. Pellet cells by centrifugation and remove supernatant.
  3. Resuspend cells in 100 µl of diluted primary antibody, prepared in Antibody Dilution Buffer at a recommended dilution or as determined via titration.
  4. Incubate for 30 min to 1 hr on ice. Protect from light.
  5. Wash by centrifugation in Antibody Dilution Buffer. Discard supernatant. Repeat.
  6. Resuspend cells in 200-500 µl of Antibody Dilution Buffer and analyze on flow cytometer.

posted June 2017

revised June 2020

Protocol Id: 1504

Specificity / Sensitivity

4-1BB/CD137/TNFRSF9 (D2Z4Y) Rabbit mAb (PE Conjugate) recognizes endogenous levels of total 4-1BB/CD137/TNFRSF9 protein.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a recombinant protein specific to the extracellular domain of human 4-1BB/CD137/TNFRSF9 protein.

Background

TNFRSF9 is a member of the tumor necrosis factor receptor superfamily (1, 2). It is also called 4-1BB or CD137 (1, 2). 4-1BB/CD137/TNFRSF9 is expressed in activated CD4+ and CD8+ T cells, natural killer cells and dendritic cells (2-5). The ligand 4-1BBL/CD137L/TNFSF9 on antigen presenting cells binds to 4-1BB/CD137/TNFRSF9 and costimulates the activation of T cells (5). The binding of agonistic antibodies to 4-1BB/CD137/TNFRSF9 also leads to costimulation for T cell activation (5). Studies have shown the effectiveness of targeting 4-1BB/CD137/TNFRSF9 by its agonistic antibodies in cancer immunotherapy (6).

  1. Sun, Y. et al. (2002) Nat Med 8, 1405-13.
  2. Wilcox, R.A. et al. (2002) J Clin Invest 109, 651-9.
  3. Lin, W. et al. (2008) Blood 112, 699-707.
  4. Wilcox, R.A. et al. (2002) J Immunol 168, 4262-7.
  5. Melero, I. et al. (1997) Nat Med 3, 682-5.
  6. Yonezawa, A. et al. (2015) Clin Cancer Res 21, 3113-20.

Pathways & Proteins

Explore pathways + proteins related to this product.

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.