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

Clone 53-6.7 is most commonly used in vivo in mice for depletion of CD8α+ cells (cytotoxic T cells), functional blocking of CD8+ T cell activity, and blocking antigen presentation via MHC class I.

Essential applications and supporting details:

• CD8+ T cell depletion: 53-6.7 induces rapid and sustained depletion of CD8α-expressing cells, enabling experimental assessment of the role of cytotoxic T cells in immune responses, tumor rejection, transplantation models, and infection.
• Blocking antigen presentation via MHC class I: The antibody binds mouse CD8α and blocks its interaction with MHC class I, interfering with antigen recognition and T cell activation.
• Inhibition of CD8+ T cell responses: By blocking CD8α, clone 53-6.7 inhibits CD8+ T cell effector functions, such as proliferation and IL-2 production, and can suppress cytotoxic activity in vivo.
• Functional studies in vivo: Used to dissect the contribution of CD8+ T cells to disease models, immune tolerance, and pathogen defense.
• Other reported uses:
  • Blocking cytotoxicity in immune assays.
  • Immunoprecipitation and functional assays (when purified for these applications).
  • Quality control as a depletion agent in various immunological experiments.

Additional relevant information:

• 53-6.7 is not recommended for immunohistochemistry on formalin-fixed paraffin sections but is validated for acetone-fixed frozen and zinc-fixed paraffin-embedded tissues.
• The antibody is available in functional-grade formulations optimized for in vivo use and tested to ensure low endotoxin and aggregation for reliable T cell depletion/blocking.

In summary, in vivo applications of clone 53-6.7 in mice center on CD8+ T cell depletion, functional blocking, and studies of T cell–dependent immunity.

Commonly Co-Used Antibodies/Proteins with 53-6.7

The 53-6.7 antibody recognizes mouse CD8α, a coreceptor expressed on cytotoxic T cells and a subset of thymocytes. In experimental immunology, it is often used in combination with other antibodies and proteins to characterize immune cell populations or to facilitate functional studies. Here are the most commonly co-used reagents in the literature:

Frequently Co-Used Antibodies

• Anti-CD4: Used alongside 53-6.7 to distinguish helper (CD4⁺) and cytotoxic (CD8⁺) T cell subsets within the broader T lymphocyte population. This is crucial for immunophenotyping in flow cytometry and immunohistochemistry.
• Anti-CD3: A pan-T cell marker that identifies all mature T cells, often paired with 53-6.7 and anti-CD4 to further delineate T cell subsets.
• Anti-CD45: The leukocyte common antigen, used to gate on all leukocytes or to distinguish T cells from other immune cell types.
• Anti-CD8b: Used to distinguish between CD8αα homodimers and CD8αβ heterodimers, providing additional resolution in studies of T cell development and function.

Additional Markers and Controls

• Isotype controls: Essential for establishing staining specificity, especially in flow cytometry and functional assays involving 53-6.7.
• Other leukocyte markers (e.g., CD19 for B cells, CD11b for myeloid cells): Occasionally co-used to exclude non-T cell populations in complex immunophenotyping panels.

Co-Used Proteins and Functional Ligands

• MHC class I molecules: The natural ligand for CD8, MHC class I is directly relevant in experiments where 53-6.7 is used to block or modulate antigen presentation and T cell activation.
• IL-2: Since 53-6.7 can inhibit T cell responses to IL-2, studies involving T cell proliferation or cytokine responses often co-assay IL-2 or use anti-IL-2 antibodies.

Summary Table

These combinations are standard in mouse immunology to characterize, isolate, and functionally manipulate CD8⁺ T cells in both in vitro and in vivo contexts. The exact panel will vary depending on the experimental question, but CD4, CD3, and CD45 are nearly universal partners for 53-6.7 in multiparameter flow cytometry and related techniques.

Clone 53-6.7 is a widely used monoclonal antibody specific for mouse CD8α, and key findings in the literature center on its diverse experimental uses and mechanistic properties.

• Functional Blocking and Depletion: Clone 53-6.7 can block antigen presentation via MHC class I and inhibit T cell responses to interleukin-2 (IL-2). It is frequently used to functionally deplete or block CD8α⁺ T cells in vivo and in vitro, thereby allowing studies of CD8⁺ T cell roles in immunity or disease models.
• Detection and Sorting: The antibody is a primary tool in flow cytometry, immunoprecipitation, and immunohistology to identify, quantify, or isolate CD8α⁺ T cell populations in mouse tissues.
• Mechanistic Insights: CD8, recognized by 53-6.7, acts as a co-receptor binding to MHC class I and is essential for T cell activation and antigen-specific responses. Blocking CD8 can directly impact cytotoxicity and T cell proliferation.
• Experimental Notes:
  • The antibody is not recommended for formalin-fixed paraffin sections but is suitable for frozen or zinc-fixed tissues.
  • Functional grade preparations are advised for in vivo experiments to minimize confounding effects of preservatives or endotoxin.
  • The antibody recognizes both CD8αα and CD8αβ forms, thus labeling a variety of immune cells beyond conventional CD8αβ cytotoxic T cells, including specific dendritic cells and intraepithelial lymphocytes.

Summary Table: Main Uses and Effects

Overall, clone 53-6.7 is considered a key reagent for mouse immunology research, essential for studying CD8 T cell biology, immune responses, and experimental immunodepletion or blockade protocols.

The dosing regimens for clone 53-6.7, a rat anti-mouse CD8α monoclonal antibody, can vary significantly across different mouse models. This variation is influenced by several factors, including the specific experimental objective, mouse strain, route of administration, and whether the goal is acute depletion or chronic suppression of CD8+ T cells.

Factors Influencing Dosing Regimens

1. Experimental Objective: The desired outcome, such as depletion of CD8+ T cells or inhibition of T cell responses, affects the dosing schedule.
2. Mouse Strain: Different strains may require adjusted dosages due to varying immune responses.
3. Route of Administration: Intraperitoneal (i.p.) or intravenous (i.v.) routes are commonly used, with i.p. being more frequent for in vivo studies.

Typical Dosing Ranges

While specific dosing ranges for clone 53-6.7 are not universally standardized, typical applications often involve administering doses in the range of 100 μg to several hundred micrograms per mouse. However, precise dosages can depend on the specific requirements of the study and are often determined experimentally.

Applications and Considerations

• Applications: Clone 53-6.7 is used for blocking antigen presentation via MHC class I, inhibiting T cell responses to IL-2, and depleting CD8a+ cells.
• Considerations: The efficacy and potential side effects of depleting CD8+ T cells must be considered, as these cells play a crucial role in immune responses, particularly against viral infections and tumors.

In summary, the dosing regimen for clone 53-6.7 in mouse models is highly dependent on the specific experimental context, and dosages should be optimized based on preliminary studies or established protocols for similar applications.