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Purified NA/LE Mouse Anti-Human CD253
Purified NA/LE Mouse Anti-Human CD253

Flow Cytometric Analysis of TRAIL-induced Killing and RIK-2 Blocking using FITC Annexin V staining.  Jurkat T cells were left untreated (far left/first panel) or treated for 16 hours under the following conditions: Cells were incubated with 20 ng/ml of recombinant human TRAIL and 10 µg/ml anti-histidine antibody (second panel); or with 20 ng/ml of recombinant human TRAIL and 10 µg/ml anti-histidine antibody preincubated with RIK-2 antibody (third panel); or with 20 ng/ml of recombinant human TRAIL and 10 µg/ml anti-histidine antibody preincubated with mIgG1 antibody (negative control) (far right/fourth panel). Following treatments, cells were evaluated for Annexin-V staining. Cells induced to undergo apoptosis by treatment with recombinant human TRAIL gave a population of cells that was Annexin V-FITC positive (second panel, M2). Annexin V-FITC staining was blocked when cells were incubated with a mixture of recombinant human TRAIL and RIK-2 antibody (third panel, M2). Cells treated with the mixture of recombinant TRAIL and mIgG1 antibody could not block the killing (far right/fourth panel). A small number of Annexin-V positive cells in the untreated population represents a basal level of apoptosis (far left/first panel). The results indicate that clone RIK-2 can block cell mediated killing induced by recombinant human TRAIL as measured by Annexin V-FITC staining of Jurkat cells.

Flow Cytometric Analysis of TRAIL-induced Killing and RIK-2 Blocking using FITC Annexin V staining.  Jurkat T cells were left untreated (far left/first panel) or treated for 16 hours under the following conditions: Cells were incubated with 20 ng/ml of recombinant human TRAIL and 10 µg/ml anti-histidine antibody (second panel); or with 20 ng/ml of recombinant human TRAIL and 10 µg/ml anti-histidine antibody preincubated with RIK-2 antibody (third panel); or with 20 ng/ml of recombinant human TRAIL and 10 µg/ml anti-histidine antibody preincubated with mIgG1 antibody (negative control) (far right/fourth panel). Following treatments, cells were evaluated for Annexin-V staining. Cells induced to undergo apoptosis by treatment with recombinant human TRAIL gave a population of cells that was Annexin V-FITC positive (second panel, M2). Annexin V-FITC staining was blocked when cells were incubated with a mixture of recombinant human TRAIL and RIK-2 antibody (third panel, M2). Cells treated with the mixture of recombinant TRAIL and mIgG1 antibody could not block the killing (far right/fourth panel). A small number of Annexin-V positive cells in the untreated population represents a basal level of apoptosis (far left/first panel). The results indicate that clone RIK-2 can block cell mediated killing induced by recombinant human TRAIL as measured by Annexin V-FITC staining of Jurkat cells.

Product Details
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BD Pharmingen™
TNFSF10; TRAIL; APO-2L; TL2
Human (QC Testing)
Mouse IgG1
Human TRAIL Transfected Cell Line
Flow cytometry (Routinely Tested), Bioassay, Blocking (Tested During Development)
1.0 mg/ml
AB_393955
No azide/low endotoxin: Aqueous buffered solution containing no preservative, 0.2µm sterile filtered. Endotoxin level is ≤0.01 EU/µg (≤0.001 ng/µg) of protein as determined by the LAL assay.
RUO


Preparation And Storage

Store undiluted at 4°C. This preparation contains no preservatives, thus it should be handled under aseptic conditions. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
550912 Rev. 6
Antibody Details
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RIK-2

The RIK-2 monoclonal antibody specifically binds to CD253, also known as TRAIL (TNF-Related Apoptosis-Inducing Ligand) and Apo2 ligand (APO-2L). CD253 is a member of the Tumor Necrosis Factor Superfamily and is encoded by the TNFSF10 gene. CD253 is a type II membrane protein that may be expressed as a soluble as well as full-length, cell surface-associated protein. Both surface and soluble forms of TRAIL rapidly induce apoptosis in a wide range of tumor cell lines but not normal tissue. TRAIL is expressed by activated T cells, NK cells, dendritic cells, monocytes and a variety of non-lymphoid cells. TRAIL can bind to and exert apoptosis through DR4 (TRAIL-R1) and DR5 (TRAIL-R2) receptors. It can also bind to decoy receptors, including DcR1/TRID/TRAIL-R3 and DcR2/TRUNDD/TRAIL-R4, and possibly OPG/TNFRSF11B, which may serve to regulate TRAIL activity. TRAIL has been shown to be involved in T cell-mediated cytotoxicity, but its mechanism of action remains to be fully elucidated. The RIK-2 clone was selected based on its ability to block TRAIL-mediated cytotoxic activity.

550912 Rev. 6
Format Details
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NA/LE
NA/LE refers to the culture and purification methods and buffer used to produce purified antibodies with no azide and low endotoxin: Aqueous buffered solution containing no preservative, 0.2µm sterile filtered. Endotoxin level is ≤0.01 EU/µg (≤0.001 ng/µg) of protein as determined by the LAL assay.NA/LE are perfectly suited to be used in culture or in vivo (for nonhuman studies) for functional assays — blocking, neutralizing, activation or depletion — where the presence of azide may damage cells or exogenous endotoxin may signal or activate cells.
NA/LE
550912 Rev.6
Citations & References
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Development References (6)

  1. Kayagaki N, Yamaguchi N, Nakayama M, et al. Involvement of TNF-related apoptosis-inducing ligand in human CD4+ T cell-mediated cytotoxicity. J Immunol. 1999; 162(5):2639-2647. (Immunogen: Blocking). View Reference
  2. Mariani SM, Matiba B, Armandola EA, Krammer PH. Interleukin 1 beta-converting enzyme related proteases/caspases are involved in TRAIL-induced apoptosis of myeloma and leukemia cells. J Cell Biol. 1997; 137(1):221-229. (Biology). View Reference
  3. Marsters SA, Pitti RM, Donahue CJ, Ruppert S, Bauer KD, Ashkenazi A. Activation of apoptosis by Apo-2 ligand is independent of FADD but blocked by CrmA. Curr Biol. 1996; 6(6):750-752. (Biology). View Reference
  4. Pitti RM, Marsters SA, Ruppert S, Donahue CJ, Moore A, Ashkenazi A. Induction of apoptosis by Apo-2 ligand, a new member of the tumor necrosis factor cytokine family. J Biol Chem. 1996; 271(22):12687-12690. (Biology). View Reference
  5. Sheridan JP, Marsters SA, Pitti RM, et al. Control of TRAIL-induced apoptosis by a family of signaling and decoy receptors. Science. 1997; 277(5327):818-821. (Biology). View Reference
  6. Wiley SR, Schooley K, Smolak PJ, et al. Identification and characterization of a new member of the TNF family that induces apoptosis. Immunity. 1995; 3(6):673-682. (Biology). View Reference
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550912 Rev. 6

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For Research Use Only. Not for use in diagnostic or therapeutic procedures.