(9 references)-
- 抗原 See all CD8 alpha (CD8A) products
- CD8 alpha (CD8A) (CD8a Molecule (CD8A))
- 适用
- 小鼠
- 宿主
- 大鼠
- 克隆类型
- 单克隆
- 标记
- Magnetic Particles
- 应用范围
- Separation (Sep)
- 品牌
- BD IMag™
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- 实验流程
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1. Prepare a single-cell suspension from the lymphoid tissue of interest according to standard laboratory procedures. Remove clumps of cells and/or debris by passing the suspension through a 70-µm nylon cell strainer.
2. Dilute BD IMag™ Buffer (10X) (Cat. No. 552362) 1:10 with sterile distilled water or prepare 1X BD IMag™ buffer by supplementing Phosphate Buffered Saline with 0.5% BSA, 2 mM EDTA, and 0.09% sodium azide. Place on ice. Although our experience indicates that the use of Mouse BD Fc Block™ purified anti-mouse CD16/CD32 mAb 2.4G2 (Cat. No. 553141/553142) is not required for optimal cell separation, some laboratories may want to use it in their studies. If adding Mouse BD Fc Block, proceed to Step 3. If not adding Mouse BD Fc Block, proceed to Step 4.
3. Add Mouse BD Fc Block at 0.25 µg/10^6 cells, and incubate on ice for 15 minutes.
4. Wash cells with at least an equal volume of 1X BD IMag buffer, and carefully aspirate all the supernatant.
5. Vortex the BD™ IMag anti-mouse CD8a Particles - DM thoroughly, and add 50 µl of particles for every 10^7 total cells.
6. MIX THOROUGHLY. Refrigerate at 6°C - 12°C for 30 minutes.
7. Bring the BD IMag-particle labeling volume up to 1 - 8 x 10^7 cells/ml with 1X BD IMag™ buffer, and immediately place the tube on the BD IMagnet™. Incubate at room temperature for 6 - 8 minutes.
8. With the tube on the BD IMagnet™, carefully aspirate off the supernatant. This supernatant contains the negative fraction.
9. Remove the tube from the BD IMagnet™, and add 1X BD IMag buffer to the same volume as in Step 7. Gently resuspend cells by pipetting briefly, and return the tube to the BD IMagnet™ for another 2 - 4 minutes.
10. With the tube on the BD IMagnet™, carefully aspirate off the supernatant and discard.
11. Repeat Steps 9 and 10.
12. After the final wash step, resuspend the positive fraction in an appropriate buffer and at an appropriate concentration for further analysis.
NOTE: Avoid nonspecific labeling by working quickly and adhering to recommended incubation times. - 限制
- 仅限研究用
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- 状态
- Liquid
- 缓冲液
- Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
- 储存液
- Sodium azide
- 注意事项
- This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
- 储存条件
- 4 °C
- 储存方法
- Store undiluted at 4° C.
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: "Development of CD8alpha-positive dendritic cells from a common myeloid progenitor." in: Science (New York, N.Y.), Vol. 290, Issue 5499, pp. 2152-4, (2000) (PubMed).
: "TAP-independent selection of CD8+ intestinal intraepithelial lymphocytes." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 156, Issue 11, pp. 4209-16, (1996) (PubMed).
: "Thymus-neuroendocrine interactions in extrathymic T cell development." in: Science (New York, N.Y.), Vol. 265, Issue 5180, pp. 1860-2, (1994) (PubMed).
: "Extrathymic differentiation of intraepithelial lymphocytes: generation of a separate and unequal T-cell repertoire?" in: Immunology today, Vol. 12, Issue 12, pp. 436-8, (1992) (PubMed).
: "The surface phenotype of dendritic cells purified from mouse thymus and spleen: investigation of the CD8 expression by a subpopulation of dendritic cells." in: The Journal of experimental medicine, Vol. 176, Issue 1, pp. 47-58, (1992) (PubMed).
: "Selective expression of CD8 alpha (Ly-2) subunit on activated thymic gamma/delta cells." in: European journal of immunology, Vol. 20, Issue 4, pp. 927-30, (1990) (PubMed).
: "Comparison of thymic and peripheral T cell Ly-2/3 antigens." in: European journal of immunology, Vol. 14, Issue 10, pp. 906-10, (1984) (PubMed).
: "Fluorescence analysis and anatomic distribution of mouse T lymphocyte subsets defined by monoclonal antibodies to the antigens Thy-1, Lyt-1, Lyt-2, and T-200." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 127, Issue 6, pp. 2594-604, (1982) (PubMed).
: "Xenogeneic monoclonal antibodies to mouse lymphoid differentiation antigens." in: Immunological reviews, Vol. 47, pp. 63-90, (1980) (PubMed).
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: "Development of CD8alpha-positive dendritic cells from a common myeloid progenitor." in: Science (New York, N.Y.), Vol. 290, Issue 5499, pp. 2152-4, (2000) (PubMed).
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- 抗原
- CD8 alpha (CD8A) (CD8a Molecule (CD8A))
- 别名
- CD8a (CD8A 产品)
- 别名
- cd8 Accessory Reagents, LOC100136450 Accessory Reagents, BB154331 Accessory Reagents, Ly-2 Accessory Reagents, Ly-35 Accessory Reagents, Ly-B Accessory Reagents, Lyt-2 Accessory Reagents, zgc:136643 Accessory Reagents, CD8 Accessory Reagents, Leu2 Accessory Reagents, MAL Accessory Reagents, p32 Accessory Reagents, RHACD8-4 Accessory Reagents, CD8a molecule Accessory Reagents, uncharacterized LOC100125537 Accessory Reagents, CD8 alpha Accessory Reagents, CD8 antigen, alpha chain Accessory Reagents, T-cell surface glycoprotein CD8 alpha chain Accessory Reagents, Cd8a Accessory Reagents, LOC100125537 Accessory Reagents, LOC100136450 Accessory Reagents, CD8A Accessory Reagents, cd8a Accessory Reagents, LOC100126556 Accessory Reagents
- 背景
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BD IMag™ anti-mouse CD8a Particles - DM are magnetic nanoparticles that have monoclonal antibody conjugated to their surfaces. These particles are optimized for the positive selection or depletion of CD8a-bearing leukocytes using the BD IMagnet™. CD8a has been reported to be expressed on most thymocytes and a subpopulation of mature T lymphocytes (e.g. MHC class I-restricted T cells, including most T suppressor/cytotoxic cells). In addition, subsets of gammadelta TCR-bearing T cells, intestinal intraepithelial lymphocytes, and dendritic cells also have been reported to express CD8a.
Leukocytes are labeled with BD IMag™ anti-mouse CD8a Particles - DM according to the Magnetic Labeling Protocol. This labeled cell suspension is then placed within the magnetc field of BD IMagnet™ (Cat. No. 552311). Labeled cells migrate toward the magnet (positive fraction), leaving the unlabeled cells in suspension so they can be drawn off (negative fraction). The tube is then removed from the magnetic field for resuspension of the positive fraction. The seperation is repeated twice to increase the purity of the positive fraction. The magnetic separation steps are diagrammed in the Separation Flow Chart. After the positive fraction is washed, the small size of the magnetic particles allows the positive fraction to be further evaluated in downstream applications such as flow cytometry.
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