Anti-Mouse CD8a (Ly 2) [Clone 53-6.7] — Purified in vivo PLATINUM™ Functional Grade

Clone 53-6.7 is most commonly used in vivo in mice for depletion of CD8a⁺ cells (cytotoxic T lymphocytes), blocking antigen presentation via MHC class I, and inhibition of CD8 T cell responses, including IL-2 production.

Key in vivo applications include:

• Depleting CD8+ T cells: Administration of clone 53-6.7 can selectively deplete CD8a⁺ T cells from peripheral blood, spleen, and lymphoid organs to study their role in immune responses or disease models.
• Functional blocking of CD8+ T cells: The antibody can block CD8α interactions, preventing cytotoxic T cell activation and thus enabling assessment of their necessity in immune processes.
• Blocking antigen presentation through MHC class I: By binding CD8a, the antibody inhibits the CD8/MHC class I interaction required for antigen recognition by cytotoxic T cells.
• Suppressing CD8-mediated T cell responses: Experimental use includes inhibition of proliferation and cytokine production (e.g., IL-2) by CD8⁺ T cells.
• Analysis of immune mechanisms: Depleting or blocking CD8+ T cells allows researchers to analyze the role of these cells in infection, cancer, autoimmunity, transplantation, and vaccine responses.

Additional applications, though less common or typically conducted in vitro, include:

• Cell isolation for adoptive transfer or functional studies.
• Immunoprecipitation and immunofluorescence to monitor depletion and verify antibody activity.

These uses are central for studies requiring mechanistic understanding of CD8+ T cells or for validating interventions that require their removal or inhibition. It is essential to use a functional grade, endotoxin-free preparation for in vivo work to avoid potential confounding immune activation.

The antibody 53-6.7, which targets mouse CD8α, is most commonly co-used with several key antibodies and proteins in immunological research. The main ones frequently used alongside 53-6.7 include:

• CD4: Marker for helper T cells—used to distinguish CD4+ and CD8+ populations.
• CD3: Pan-T cell marker—used to identify all T cells and analyze T cell subsets.
• CD45: A pan-leukocyte (white blood cell) marker—commonly used for identifying all leukocytes in a mixture.
• CD8β: Used to differentiate CD8αβ (cytotoxic T cells) from CD8αα (other subsets) T cells.

These markers are fundamental in mouse immunology panels, supporting reliable T cell subset analysis, gating strategies in flow cytometry, and functional studies of T cell responses.

Summary Table: Commonly Co-Used Antibodies with 53-6.7

Additional frequently used markers (depending on study goals):

• Isotype controls: Matched to the species and fluorochrome for quality control.
• Other lineage or activation markers: Such as CD44, CD62L for naïve/memory classification; CD25 for activation status.

Functional proteins and reagents that may appear in conjunction with 53-6.7:

• MHC class I tetramers for antigen-specific T cell identification.
• Cytokine staining (e.g., IFN-γ, TNF) for functional profiling.

Context in Literature:In flow cytometry, 53-6.7 is nearly always paired with these markers (primarily CD4, CD3, and CD45) for complete lymphocyte panel construction and functional immunology studies.

If you are designing panels or searching literature for multiplex analyses or cell sorting, these are the most common combination partners for 53-6.7.

Clone 53-6.7 is a monoclonal antibody used extensively in mouse immunology research to specifically bind the CD8α chain of the CD8 co-receptor, marking cytotoxic T lymphocytes and enabling their identification, depletion, or functional inhibition.

Key findings and established uses from scientific literature include:

• Specificity and Target: 53-6.7 binds both the CD8α (38 kDa) and CD8α' (34 kDa) chains (Ly-2/Lyt-2) of the mouse CD8 differentiation antigen, present on cytotoxic T cells across all mouse strains tested.
• Blocking and Functional Inhibition:
  • This antibody blocks antigen presentation via MHC class I, thereby interfering with antigen recognition processes by CD8+ T cells.
  • It inhibits T cell responses to IL-2 (Interleukin-2), which normally promotes T cell proliferation and survival.
  • It has been used to block cytotoxic activity of CD8+ T cells and their proliferation, helping dissect their precise role in immune responses.
• Cell Depletion: 53-6.7 is routinely employed to deplete CD8α+ cells both in vitro and in vivo, making it a key tool in studies requiring removal or functional neutralization of cytotoxic T cell populations.
• Competitive Binding: 53-6.7 competes with other anti-CD8 antibodies (e.g., clone 5H10-1) for binding to thymocytes, useful in comparative studies of CD8-targeting reagents.
• Applications:
  • Flow cytometry for quantitation and phenotyping of CD8+ T cells.
  • Immunoprecipitation of CD8a for biochemical analyses.
  • Immunohistochemical staining of frozen and zinc-fixed tissue sections (not recommended for formalin-fixed paraffin sections).
  • Spatial biology and imaging (e.g., IBEX-based multiplexed antibody panels).
• Recommended Usage: Typically less than or equal to 0.25 µg per test for flow cytometry with optimal titration required depending on the assay and sample size.

Technical and experimental findings confirm that 53-6.7 has been fundamental in studies of T cell–mediated immunity, transplantation (tracking clonal diversity and expansion of graft-specific CD8+ T cells), tumor immunology (characterizing tumor-infiltrating lymphocytes), and basic T cell biology.

In summary, clone 53-6.7 is a foundational reagent in mouse immunology for identifying, depleting, and functionally characterizing CD8+ T cells, with reliable specificity and broad application across immune, cancer, and transplantation research.

Dosing regimens for clone 53-6.7 in mice are highly variable and depend on the mouse model, experimental objective, strain, and route of administration. While no universal standard exists, most studies utilize doses within the range of 100–500 μg per mouse per injection, typically administered intraperitoneally or intravenously.

Key contextual details:

• Experimental Objective: Dosing differs if the intent is acute depletion versus chronic suppression of CD8⁺ T cells. Acute depletion often uses higher, single or initial doses, whereas chronic protocols feature repeated dosing at lower maintenance levels.
• Mouse Strain and Model: Variability arises due to different strains’ immunological responses—studies often recommend optimizing dosages for each specific strain and experimental setup.
• Regimen Examples:
  • For in vivo CD8⁺ T cell depletion, protocols commonly start with 250–500 μg per mouse, repeated every 3–7 days to maintain depletion, but optimization is needed for disease models, tumor presence, or immune activation state.
• Route of Administration: Intraperitoneal and intravenous injections are standard; intravenous administration may yield more rapid effects but could require adjusted dosing.
• Controls and Validation: Effective depletion must be confirmed via flow cytometry using appropriate controls, as published protocols recommend.

Summary table of factors influencing clone 53-6.7 dosing:

Additional notes:

• Most references recommend preliminary dose titration studies within a given model to confirm the minimum dose required for effective CD8⁺ T cell depletion, minimizing off-target effects and immunogenicity.
• Some sources mention that for flow cytometry or cell sorting, as little as ≤0.25 μg per 10⁶ cells is used for labeling, not depletion.
• When compared to other clones (e.g., 2.43 or YTS 169.4), 53-6.7 performs similarly but should be selected according to pre-existing model validation and compatibility with desired endpoints.

In conclusion, clone 53-6.7 dosing regimens must be customized to each mouse model and experimental goal, with typical depletion doses ranging from 100–500 μg per mouse, repeated as needed, and validated by flow cytometry.