Anti-Mouse CD8b.2 [Clone 53-5.8] — Purified in vivo GOLD™ Functional Grade

In Vivo Applications of Clone 53-5.8 in Mice

The monoclonal antibody clone 53-5.8, specific for mouse CD8β (Lyt 3.2), is a widely used reagent in immunological research for in vivo applications using mice. Its primary function is to target and deplete CD8+ T cells, which are crucial for understanding immune responses, disease mechanisms, and therapeutic interventions.

Key In Vivo Uses

• CD8+ T Cell Depletion: The most common and well-characterized application of clone 53-5.8 is the selective depletion of CD8+ T cells in living mice. When administered in vivo, this antibody effectively removes CD8+ T cells while sparing other immune cell populations, making it a valuable tool for studying the role of these cells in immune responses, infection, cancer, autoimmunity, and transplantation.
• Functional Studies in Immunity: Researchers use clone 53-5.8 to investigate the consequences of CD8+ T cell absence in various disease models. For example, it has been employed to study how CD8+ T cell depletion affects tumor immunity, viral infection outcomes, and transplant rejection.
• Comparative Depletion Strategies: Studies have shown that clone 53-5.8 (anti-CD8β) is more effective at depleting CD8+ T cells than anti-CD8α antibodies like clone 53-6.7. The surviving CD8+ T cell populations after depletion with 53-5.8 exhibit distinct phenotypic and functional differences compared to those surviving anti-CD8α treatment, which is important for interpreting experimental results.
• Mechanistic Insights: The use of 53-5.8 has revealed that CD8+ T cells surviving anti-CD8β depletion may have enhanced cytotoxic function compared to those surviving anti-CD8α depletion, which could influence the outcome of immune responses.
• Model-Specific Considerations: Clone 53-5.8 is particularly effective in mouse strains expressing the Ly-3.2 allele of CD8β, and its specificity for this allele makes it less reactive with Ly-3.1-expressing strains.

Typical Protocols

• Dosage: A standard in vivo dose is 200 μg per mouse or 10 mg/kg, administered intraperitoneally.
• Timing: Antibody administration can occur before, during, or after experimental manipulation, depending on the research question (e.g., pre-tumor implantation for cancer studies or pre-infection for viral models).
• Combination Studies: Clone 53-5.8 is sometimes used alongside other depleting antibodies (e.g., anti-CD4, anti-CD25) to investigate the roles of multiple immune cell subsets.

Limitations and Caveats

• Residual CD8+ T Cells: While clone 53-5.8 depletes most CD8+ T cells, a small population may survive and remain functional, which can affect experimental interpretations.
• Reduced CD8 Staining: The antibody can induce substantial internalization of CD8, making flow cytometric detection of surviving CD8+ T cells challenging without careful gating.
• Strain Specificity: Effectiveness is greatest in mice expressing the Ly-3.2 allele of CD8β.

Summary Table

Clone 53-5.8 is a cornerstone reagent for in vivo CD8+ T cell depletion in mice, enabling precise dissection of the roles these cells play in health and disease. Its use requires careful experimental design and interpretation due to potential residual cell activity and technical detection challenges.

The 53-5.8 antibody, which targets mouse CD8b.2, is commonly used in conjunction with other antibodies or proteins for various applications, particularly in immunological studies. Here are some of the other commonly used antibodies or proteins:

• Anti-CD8α: Antibodies like 53-6.7 or 53-6.72 are often used alongside 53-5.8 for comprehensive T cell characterization, as they target different parts of the CD8 complex.
• Anti-CD3: This marker is used to identify T cells in general, often in combination with CD8 or CD4 antibodies for subset identification.
• Anti-CD4: Used for identifying helper T cells, these antibodies are frequently paired with CD8 antibodies to distinguish between T cell subsets.
• Anti-CD45: Pan-leukocyte markers like anti-CD45 are used to identify leukocytes broadly, often in conjunction with more specific markers like CD8.

These combinations allow researchers to study T cell subsets, their roles in immune responses, and the effects of CD8+ T cell depletion in experimental models.

Clone 53-5.8 is a monoclonal antibody targeting mouse CD8β and is widely cited in scientific literature as a reliable and potent tool for CD8+ T cell depletion in vivo. Key findings from its use include:

• High Depletion Efficiency: Clone 53-5.8 is considered more effective than anti-CD8α antibodies at depleting CD8+ T cells in mice, making it the preferred reagent for studies requiring ablation of this cell subset.

• Disease Model Impact:

  • In malaria models, depletion of CD8+ T cells with 53-5.8 confers protection against cerebral complications and reduces pulmonary vascular leakage without altering parasite burden, implicating CD8+ T cells in disease pathology.
  • In cancer models, anti-CD8β (clone 53-5.8) treatment impairs CD8+ T cell-mediated antitumor activity, confirming their essential role in immunosurveillance and tumor rejection.

• Effect on CD8+ T Cell Function: Surviving CD8+ T cells after partial depletion may display altered functions, such as changes in activation markers or cytotoxic potential, highlighting the antibody's impact beyond simple numeric reduction.

• Immunology Research Utility: Frequently used in adoptive transfer, differentiation, and functional studies to discriminate CD8β+ subpopulations or to dissect specific T cell–mediated mechanisms in infection, autoimmunity, transplantation, and cancer models.

• Comparison with Other Clones: Findings validated using clone 53-5.8 are often cross-checked against other CD8β-targeting clones (e.g., H35-17.2), with similar results—supporting the clone’s reliability for phenotype and depletion studies.

• Mechanistic Insights:

  • Its use has revealed cell-intrinsic and extrinsic requirements for CD8+ T cell differentiation and trafficking, such as the requirement of CD103 or E-cadherin interactions, by enabling selective depletion or labeling of CD8β+ cells in complex immunological settings.
  • The clone facilitates the analysis of regulatory interactions during T cell development (e.g., coreceptor-LCK interactions).

In summary, clone 53-5.8 is highly regarded for efficient and specific CD8+ T cell depletion, and its use has provided crucial insights into the roles of these cells in infection, immunity, and disease models.

Dosing regimens for clone 53-5.8, an anti-mouse CD8β monoclonal antibody, can vary depending on the specific mouse model and experimental design. Here are some general insights into how these regimens might differ across different models:

1. Dosage: The typical dosage for clone 53-5.8 is often around 5 mg/kg when administered intraperitoneally (i.p.) in various studies. However, some studies use a much lower dose of 20 μg per mouse, also administered i.p., for specific experimental needs.

2. Administration Schedule: The frequency of administration can vary. For CD8+ T cell depletion studies, it might be administered on the day before and the day of tumor implantation, followed by weekly injections. In other scenarios, it could be administered weekly from day 0 of the experiment.

3. Applications: Clone 53-5.8 is used for CD8+ T cell depletion in diverse models, including tumor immunity, viral infections, and autoimmune diseases. The dosing regimen can be tailored to the specific requirements of each model, such as the timing of cell depletion relative to other treatments or interventions.

4. Species and Strain: While the antibody is often used in mice, the specific dosing may depend on the strain or breeding conditions, which can influence the response to the treatment. However, detailed strain-specific variations are not frequently reported in the literature.

Overall, while the basic dosing regimen for clone 53-5.8 tends to involve intraperitoneal injections of 5 mg/kg or similar dosages, the specific details can vary based on experimental objectives and the mouse model used.