CD226 antibody | DX11
Low endotoxin Mouse anti Human CD226 antibody, clone DX11 (MCA2257EL) used for blocking DNAM-1 on human NK cells to evaluate the role of DNAM-1 in NK cell mediated NK cytotoxicity.
Image caption:
NO exposed SKO-007(J3) cells enhances NK cell-mediated cytotoxicity. A) NK cells prepared from PBMCs of healthy donors, were incubated with SKO-007(J3) cells, untreated or treated with DETA-NO for 48 h, and used as target cells in a degranulation assay. The assay was performed at the effector:target (E:T) ratio of 2.5:1. After 2 hours at 37°C, cells were stained with anti-CD56, anti-CD3 and anti-CD107a mAbs. Cell surface expression of CD107a was analyzed on CD56+CD3− cells. In order to evaluate the role of DNAM-1, the assay was performed in parallel treating NK cells with blocking anti-DNAM-1 antibody. Results are representative of one out of three independent experiments. B) The MFI of CD107 were calculated based on at least three independent experiments and evaluated by paired Student t test (*P <0.05). Histograms represent the MFI with specific mAb subtracted from the MFI value of isotype control. C) NK cells isolated from PBMCs of healthy donors were incubated with SKO-007(J3) cells, untreated or treated with DETA-NO for 48 h as described above, and used as target cells in a standard 4-hour chromium-release assay. The percentage of specific lysis was calculated by counting an aliquot of supernatant and using the formula: 100 x [(sample release - spontaneous release)/total release - spontaneous release)]. All determinations were made in triplicate and E:T ratios ranged from 10:1 to 1:1, as indicated. Data represent the mean (n = 3 experiments, *P <0.05).
From: Fionda C, Abruzzese M, Zingoni A, Soriani A, Ricci B, Molfetta R, Paolini R, Santoni A, Cippitelli M.
Nitric oxide donors increase PVR/CD155 DNAM-1 ligand expression in multiple myeloma cells: role of DNA damage response activation.
BMC Cancer. 2015 Jan 22;15(1):17.
This image is from an open access article distributed under terms of a Creative Commons Attribution License.
Mouse anti Human CD266 antibody, clone DX11 (MCA2257) used to evaluate DNAM-1 (CD226) expression on HS5 cells by flow cytometry.
Image caption:
CD155 promotes Multiple Myeloma (MM) cell binding to bone marrow stromal cells (BMSCs). (A) CD155 knock-down in the Multiple Myeloma cell line ARK was achieved by means of lentiviral-mediated shRNA targeting (pLKO vector, MISSION™ Sigma-Aldrich). CD155 surface expression was analyzed by FACS in cells infected with shCD155 (code SHCLND-NM_006505, sequence TRCN0000062911) (empty red histogram) or non-targeting shRNA (Mock, filled gray histogram) by mean of a PE-conjugated anti-CD155 monoclonal Ab (clone SKII.4, Biolegend). Isotype control staining is also shown (Ctrl Ig, empty black histogram). Data from one representative experiment out of three independent experiments is shown. (B, left panel) CD155-silenced or mock-infected ARK cells were labeled with the red fluorescent dye PKH26 and left to adhere to a monolayer of HS5 bone marrow-derived stromal cells for 2 h at 37°C. Cells were washed three times with PBS and fixed. Adherent cells were visualized with IX73 microscope equipped with a 10 X objective (Olympus). An overlay image mixing red fluorescence and brightfield is shown, scale bar: 50μm. (B, right panel) Adherent cells were quantified with FIJI software. Means ± SD of 15 randomly acquired fields from two independent experiments are shown. ** p <0.01, Unpaired t-test. (C) HS5 stromal cell line were stained for Nectin3 (clone N3.12.4, Millipore), DNAM-1 (clone DX11, Serotec), CD96 (clone NK92.39, Biolegend) and TIGIT (clone A15153G, Biolegend) (empty red histograms) or with isotype matched Ctrl Ig (black histograms) followed by an APC-conjugated goat anti-mouse Ab (Jackson Laboratories) and analyzed by flow cytometry. Data from one representative experiment out of three independent experiments is shown.
From: Molfetta, R.; Zitti, B.; Lecce, M.; Milito, N.D.; Stabile, H.; Fionda, C.; Cippitelli, M.; Gismondi, A.; Santoni, A.; Paolini, R.
CD155: A Multi-Functional Molecule in Tumor Progression.
Int J Mol Sci 2020, 21, 922.
doi: 10.3390/ijms21030922
This image is from an open access article distributed under terms of a Creative Commons Attribution License.
Mouse anti Human CD226 antibody, clone DX11 (MCA2257) used to treat NK cells in a Chromium release assay.
Image caption:
Inhibition of PVR SUMOylation increases MM cell susceptibility to NK cell mediated lysis. Primary cultured NK cells were pre-treated for 20 minutes at RT with anti-DNAM1 neutralizing mAb or with anti-CD56 (Ctrl mAb) and were used as effector cells in a 4 hours 51Cr release assay (a) or in a CD107a degranulation assay (b) toward shUBC9 infected or mock infected cells. (a) NK cells were incubated with target cells at the indicated E:T ratios. The percentage of specific lysis (left panel) and Lytic Units (right panel), calculated as described in Materials and Methods, from three independent experiments (mean ±SD) are shown. *p <0.05, **p <0.01, One-way ANOVA. (b) After 4 hours of co-culture cells were stained with CD107a-APC, CD56-PE, CD3-FITC and CD138-PerCP-CY5.5 to gate the CD56+ CD3− CD138− NK cell population, and the analysis was performed with FACSCanto (BD Biosciences). Left panels: the percentage of CD107a+ NK cells is shown and is representative of three independent experiments. Right panel: Means ±SD of three independent experiments are shown. *p <0.05, **p <0.01, Two-way ANOVA.
From: Zitti, B., Molfetta, R., Fionda, C. et al.
Innate immune activating ligand SUMOylation affects tumor cell recognition by NK cells.
Sci Rep 7, 10445 (2017).doi: 10.1038/s41598-017-10403-0.
This image is from an open access article distributed under terms of a Creative Commons Attribution License.
Mouse anti Human CD226 antibody, clone DX11 (MCA2257) used to treat NK cells prior to a degranulation assay on SKO-007(J3) target cells.
Image caption:
BMSCs enhance the ability of MM cells to stimulate NK cell degranulation by promoting DNAM-1 recognition. NK cells derived from PBMCs of healthy donors were incubated with SKO-007(J3) cells, untreated or cultured for 72 h with BMSC-CM and used as target cells in a degranulation assay. The assay was performed at the effector-target (E/T) ratio of 2.5:1. Cell surface expression of CD107a was analysed on CD3−/CD138−/CD56⁺ NK cells. To evaluate the role of DNAM-1, the assay was performed in parallel treating NK cells with blocking anti–DNAM-1 or anti-IgG mAbs used as control. Results are expressed as the percentage of CD107a+ cells. A representative experiment is shown. (B) Histogram represents the mean ± SD from three independent experiments (* p < 0.05, ANOVA).
From: Mekhloufi A, Kosta A, Stabile H, Molfetta R, Zingoni A, Soriani A, Cippitelli M, Paolini R, Gismondi A, Ricciardi MR, Petrucci MT, Masuelli L, Caracciolo G, Palchetti S, Santoni A, Fionda C.
Bone Marrow Stromal Cell-Derived IL-8 Upregulates PVR Expression on Multiple Myeloma Cells via NF-kB Transcription Factor.
Cancers (Basel). 2020 Feb 13;12(2):440.
doi: 10.3390/cancers12020440.
This image is from an open access article distributed under terms of a Creative Commons Attribution License.
Mouse anti Human CD226
- Product Type
- Monoclonal Antibody
- Clone
- DX11
- Isotype
- IgG1
- Specificity
- CD226
Antibody Quality and Performance Guarantee
| Mouse anti Human CD226 antibody, clone DX11 recognizes human CD226, a ~65 kDa glycoprotein, also known as DNAM1 (DNAX accessory molecule-1). CD226 is broadly expressed on T-cells, NK cells, platelets, monocytes and a subset of B cells. CD226 is also expressed by a subset of CD3 positive thymocytes. Mouse anti Human CD226 antibody, clone DX11 is reported to inhibit T- and NK cell mediated cytotoxicity against tumor cell targets and to block TNF alpha and IFN gamma secretion by alloantigen-specific T-cells (Kojima et al. 2003). |
- Target Species
- Human
- Product Form
- Purified IgG - liquid
- Preparation
- Purified IgG prepared by affinity chromatography on Protein A from tissue culture supernatant
- Buffer Solution
- Phosphate buffered saline
- Preservative Stabilisers
- 0.09% sodium azide (NaN3)
- Carrier Free
- Yes
- Immunogen
- Human cytotoxic T lymphocyte clone
- Approx. Protein Concentrations
- IgG concentration 1.0 mg/ml
- Fusion Partners
- Spleen cells from immunized BALB/c mouse were fused with cells of the Sp2/0 myeloma cell line
- Regulatory
- For research purposes only
- Guarantee
- 12 months from date of despatch
This product is shipped at ambient temperature. It is recommended to aliquot and store at -20°C on receipt. When thawed, aliquot the sample as needed. Keep aliquots at 2-8°C for short term use (up to 4 weeks) and store the remaining aliquots at -20°C.
Avoid repeated freezing and thawing as this may denature the antibody. Storage in frost-free freezers is not recommended.
Avoid repeated freezing and thawing as this may denature the antibody. Storage in frost-free freezers is not recommended.
This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit the antibody protocols page.
| Application Name | Verified | Min Dilution | Max Dilution |
|---|---|---|---|
| Flow Cytometry |  |
1/50 Pack Size: 0.2 mg 1/25 Pack Size: 0.1 mg |
1/200 Pack Size: 0.2 mg 1/100 Pack Size: 0.1 mg |
| Immunoprecipitation | |
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
- Flow Cytometry
- Use 10μl of the suggested working dilution to label 1 x 106 cells in 100μl
| Description | Product Code | Applications | Pack Size | List Price | Your Price | Quantity | |
|---|---|---|---|---|---|---|---|
| Mouse IgG1 Negative Control | MCA928 | F | 100 Tests |
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| List Price | Your Price | ||||||
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| Description | Mouse IgG1 Negative Control | ||||||
References for CD226 antibody
-
Shibuya, A. et al. (1996) DNAM-1, a novel adhesion molecule involved in the cytolytic function of T lymphocytes.
Immunity. 4 (6): 573-81. -
Shibuya, A. et al. (1998) Protein kinase C is involved in the regulation of both signaling and adhesion mediated by DNAX accessory molecule-1 receptor.
J Immunol. 161 (4): 1671-6. -
Kojima, H. et al. (2003) CD226 mediates platelet and megakaryocytic cell adhesion to vascular endothelial cells.
J Biol Chem. 278 (38): 36748-53. -
Manes, T.D. and Pober, J.S. (2011) Identification of Endothelial Cell Junctional Proteins and Lymphocyte Receptors Involved in Transendothelial Migration of Human Effector Memory CD4+ T Cells.
J Immunol. 186: 1763-8. -
Ardolino, M. et al. (2011) DNAM-1 ligand expression on Ag-stimulated T lymphocytes is mediated by ROS-dependent activation of DNA-damage response: relevance for NK-T cell interaction.
Blood. 117: 4778-86. -
Soriani, A. et al. (2009) ATM-ATR-dependent up-regulation of DNAM-1 and NKG2D ligands on multiple myeloma cells by therapeutic agents results in enhanced NK-cell susceptibility and is associated with a senescent phenotype.
Blood. 113: 3503-11. -
Fionda, C. et al. (2009) Heat shock protein-90 inhibitors increase MHC class I-related chain A and B ligand expression on multiple myeloma cells and their ability to trigger NK cell degranulation.
J Immunol. 183 (7): 4385-94. -
Matusali, G. et al. (2012) The Human Immunodeficiency Virus Type 1 Nef and Vpu Proteins Downregulate the Natural Killer Cell-Activating Ligand PVR.
J Virol. 86: 4496-504.
View The Latest Product References
-
Fionda, C. et al. (2015) Nitric oxide donors increase PVR/CD155 DNAM-1 ligand expression in multiple myeloma cells: role of DNA damage response activation.
BMC Cancer. 15 (1): 17. -
Molfetta, R. et al. (2019) The Ubiquitin-proteasome pathway regulates Nectin2/CD112 expression and impairs NK cell recognition and killing.
Eur J Immunol. 49 (6): 873-83. -
Vulpis, E. et al. (2022) Impact on NK cell functions of acute versus chronic exposure to extracellular vesicle-associated MICA: Dual role in cancer immunosurveillance.
J Extracell Vesicles. 11 (1): e12176. -
Molfetta, R. et al. (2020) CD155: A Multi-Functional Molecule in Tumor Progression.
Int J Mol Sci. 21 (3): 922. -
Zitti, B. et al. (2017) Innate immune activating ligand SUMOylation affects tumor cell recognition by NK cells.
Sci Rep. 7 (1): 10445. -
Mekhloufi, A. et al. (2020) Bone Marrow Stromal Cell-Derived IL-8 Upregulates PVR Expression on Multiple Myeloma Cells via NF-kB Transcription Factor.
Cancers (Basel). 12 (2): 440.
- Synonyms
- DNAM-1
- RRID
- AB_324895
- UniProt
- Q15762
- Entrez Gene
- CD226
- GO Terms
- GO:0002860 positive regulation of natural killer cell mediated cytotoxicity directed against tumor cell target
- GO:0002891 positive regulation of immunoglobulin mediated immune response
- GO:0007155 cell adhesion
- GO:0050839 cell adhesion molecule binding
- GO:0004872 receptor activity
- GO:0005887 integral to plasma membrane
- GO:0005178 integrin binding
- GO:0008037 cell recognition
- GO:0009986 cell surface
- View More GO Terms
- GO:0019901 protein kinase binding
- GO:0033005 positive regulation of mast cell activation
- GO:0045121 membrane raft
- GO:0060369 positive regulation of Fc receptor mediated stimulatory signaling pathway
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