- 53-6.7 (See other available formats)
- Regulatory Status
- Other Names
- T8, Lyt2, Ly-2
- Rat IgG2a, κ
- Ave. Rating
- Submit a Review
- Product Citations
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CD8, also known as Lyt-2, Ly-2, or T8, consists of disulfide-linked α and β chains that form the α(CD8a)/β(CD8b) heterodimer and α/α homodimer. CD8a is a 34 kD protein that belongs to the immunoglobulin family. The CD8 α/β heterodimer is expressed on the surface of most thymocytes and a subset of mature TCR α/β T cells. CD8 expression on mature T cells is non-overlapping with CD4. The CD8 α/α homodimer is expressed on a subset of γ/δ TCR-bearing T cells, NK cells, intestinal intraepithelial lymphocytes, and lymphoid dendritic cells. CD8 is an antigen co-receptor on T cells that interacts with MHC class I on antigen-presenting cells or epithelial cells. CD8 promotes T cell activation through its association with the TCR complex and protein tyrosine kinase lck.Product Details
- Verified Reactivity
- Antibody Type
- Host Species
- Mouse thymus or spleen
- Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide.
- The antibody was purified by affinity chromatography and conjugated with Alexa Fluor® 647 under optimal conditions.
- 0.5 mg/mL
- Storage & Handling
- The antibody solution should be stored undiluted between 2°C and 8°C, and protected from prolonged exposure to light. Do not freeze.
FC - Quality tested
IHC-F, 3D IHC - Verified
SB - Reported in the literature, not verified in house
- Recommended Usage
Each lot of this antibody is quality control tested by immunofluorescent staining with flow cytometric analysis. For flow cytometric staining, the suggested use of this reagent is ≤ 0.25 µg per million cells in 100 µL volume. For immunohistochemistry on frozen tissue sections, a concentration range of 2.5 - 5.0 μg/mL is suggested. For 3D immunohistochemistry on formalin-fixed tissues, a concentration of 5.0 µg/mL is suggested. It is recommended that the reagent be titrated for optimal performance for each application.
* Alexa Fluor® 647 has a maximum emission of 668 nm when it is excited at 633nm / 635nm.
Alexa Fluor® and Pacific Blue™ are trademarks of Life Technologies Corporation.
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- Excitation Laser
Red Laser (633 nm)
- Application Notes
Clone 53-6.7 antibody competes with clone 5H10-1 antibody for binding to thymocytes3. The 53-6.7 antibody has been reported to block antigen presentation via MHC class I and inhibit T cell responses to IL-2. This antibody has also been used for depletion of CD8a+ cells. Additional reported applications (for the relevant formats) include: immunoprecipitation1,3, in vivo and in vitro cell depletion2,10,15, inhibition of CD8 T cell proliferation3, blocking of cytotoxicity3,4, immunohistochemical staining5,6 of acetone-fixed frozen sections and zinc-fixed paraffin-embedded sections, and spatial biology (IBEX)29,30. Clone 53-6.7 is not recommended for immunohistochemistry of formalin-fixed paraffin sections. The Ultra-LEAF™ purified antibody (Endotoxin < 0.01 EU/µg, Azide-Free, 0.2 µm filtered) is recommended for functional assays or in vivo studies (Cat No. 100746).
- Additional Product Notes
Iterative Bleaching Extended multi-pleXity (IBEX) is a fluorescent imaging technique capable of highly-multiplexed spatial analysis. The method relies on cyclical bleaching of panels of fluorescent antibodies in order to image and analyze many markers over multiple cycles of staining, imaging, and, bleaching. It is a community-developed open-access method developed by the Center for Advanced Tissue Imaging (CAT-I) in the National Institute of Allergy and Infectious Diseases (NIAID, NIH).
- Application References
- Ledbetter JA, et al. 1979. Immunol. Rev. 47:63. (IHC, IP)
- Hathcock KS. 1991. Current Protocols in Immunology. 3.4.1. (Deplete)
- Takahashi K, et al. 1992. P. Natl. Acad. Sci. USA 89:5557. (Block, IP)
- Ledbetter JA, et al. 1981. J. Exp. Med. 153:1503. (Block)
- Hata H, et al. 2004. J. Clin. Invest. 114:582. (IHC)
- Fan WY, et al. 2001. Exp. Biol. Med. 226:1045. (IHC)
- Shih FF, et al. 2006. J. Immunol. 176:3438. (FC)
- Kamimura D, et al. 2006. J. Immunol. 177:306.
- Bouwer HGA, et al. 2006. P. Natl. Acad. Sci. USA 103:5102. (FC, Deplete)
- Kao C, et al. 2005. Int. Immunol. 17:1607. PubMed
- Ko SY, et al. 2005. J. Immunol. 175:3309. (FC) PubMed
- Rasmussen JW, et al. 2006. Infect. Immun. 74:6590. PubMed
- Lee CH, et al. 2009. Clin. Cancer Res. PubMed
- Geiben-Lynn R, et al. 2008. Blood 112:4585. (Deplete) PubMed
- Kingeter LM, et al. 2008. J. Immunol. 181:6244. PubMed
- Guo Y, et al. 2008. Blood 112:480. PubMed
- Andrews DM, et al. 2008. J. Virol. 82:4931. PubMed
- Britschqui MR, et al. 2008. J. Immunol. 181:7681. PubMed
- Kenna TJ, et al. 2008. Blood 111:2091. PubMed
- Jordan JM, et al. 2008. Infect. Immun. 76:3717. PubMed
- Todd DJ, et al. 2009. J. Exp. Med. 206:2151. PubMed
- Bankoti J, et al. 2010. Toxicol. Sci. 115:422. (FC) PubMed
- Medyouf H, et al. 2010. Blood 115:1175. PubMed
- Riedl P, et al. 2009. J. Immunol. 183:370. PubMed
- Apte SH, et al. 2010. J. Immunol. 185:998. PubMed
- Bankoti J, et al. 2010. Toxicol. Sci. 115:422. (FC) PubMed
- del Rio ML, et al. 2011. Transpl. Int. 24:501. (FC) PubMed
- Cui L, et al. 2015. J Control Release. 206:220. PubMed
- Radtke AJ, et al. 2020. Proc Natl Acad Sci U S A. 117:33455-65. (SB) PubMed
- Radtke AJ, et al. 2022. Nat Protoc. 17:378-401. (SB) PubMed
- Product Citations
AB_493424 (BioLegend Cat. No. 100727)
AB_389326 (BioLegend Cat. No. 100724)
- Ig superfamily, CD8α chain, 34 kD
Most thymocytes, T cell subset, some NK cells, lymphoid dendritic cells
- Co-receptor for TCR
- MHC class I molecule
- Antigen References
1. Barclay A, et al. 1997. The Leukocyte Antigen FactsBook Academic Press.
2. Zamoyska R. 1994. Immunity 1:243.
3. Ellmeier W, et al. 1999. Annu. Rev. Immunol. 17:523.
- Gene ID
- 12525 View all products for this Gene ID
- View information about CD8a on UniProt.org
- If an antibody clone has been previously successfully used in IBEX in one fluorescent format, will other antibody formats work as well?
It’s likely that other fluorophore conjugates to the same antibody clone will also be compatible with IBEX using the same sample fixation procedure. Ultimately a directly conjugated antibody’s utility in fluorescent imaging and IBEX may be specific to the sample and microscope being used in the experiment. Some antibody clone conjugates may perform better than others due to performance differences in non-specific binding, fluorophore brightness, and other biochemical properties unique to that conjugate.
- Will antibodies my lab is already using for fluorescent or chromogenic IHC work in IBEX?
Fundamentally, IBEX as a technique that works much in the same way as single antibody panels or single marker IF/IHC. If you’re already successfully using an antibody clone on a sample of interest, it is likely that clone will have utility in IBEX. It is expected some optimization and testing of different antibody fluorophore conjugates will be required to find a suitable format; however, legacy microscopy techniques like chromogenic IHC on fixed or frozen tissue is an excellent place to start looking for useful antibodies.
- Are other fluorophores compatible with IBEX?
Over 18 fluorescent formats have been screened for use in IBEX, however, it is likely that other fluorophores are able to be rapidly bleached in IBEX. If a fluorophore format is already suitable for your imaging platform it can be tested for compatibility in IBEX.
- The same antibody works in one tissue type but not another. What is happening?
Differences in tissue properties may impact both the ability of an antibody to bind its target specifically and impact the ability of a specific fluorophore conjugate to overcome the background fluorescent signal in a given tissue. Secondary stains, as well as testing multiple fluorescent conjugates of the same clone, may help to troubleshoot challenging targets or tissues. Using a reference control tissue may also give confidence in the specificity of your staining.
- How can I be sure the staining I’m seeing in my tissue is real?
In general, best practices for validating an antibody in traditional chromogenic or fluorescent IHC are applicable to IBEX. Please reference the Nature Methods review on antibody based multiplexed imaging for resources on validating antibodies for IBEX.
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TotalSeq™-C0002 anti-mouse CD8a
TotalSeq™-B0002 anti-mouse CD8a
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PE/Fire™ 640 anti-mouse CD8a
PE/Fire™ 700 anti-mouse CD8a
Spark Blue™ 574 anti-mouse CD8a Antibody
Spark Violet™ 423 anti-mouse CD8a Antibody
Spark UV™ 387 anti-mouse CD8a
APC/Cyanine7 anti-mouse CD8a
Alexa Fluor® 488 anti-mouse CD8a
Alexa Fluor® 647 anti-mouse CD8a
Pacific Blue™ anti-mouse CD8a
Alexa Fluor® 700 anti-mouse CD8a
PerCP/Cyanine5.5 anti-mouse CD8a
PerCP anti-mouse CD8a
Brilliant Violet 421™ anti-mouse CD8a
Brilliant Violet 570™ anti-mouse CD8a
Brilliant Violet 650™ anti-mouse CD8a
Brilliant Violet 605™ anti-mouse CD8a
Ultra-LEAF™ Purified anti-mouse CD8a
Brilliant Violet 711™ anti-mouse CD8a
Brilliant Violet 785™ anti-mouse CD8a
Brilliant Violet 510™ anti-mouse CD8a
Purified anti-mouse CD8a (Maxpar® Ready)
Alexa Fluor® 594 anti-mouse CD8a