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

The clone 53-5.8 is an anti-mouse CD8? (Lyt 3.2) monoclonal antibody that serves as a powerful tool for CD8+ T cell depletion in in vivo mouse studies. This rat IgG1 antibody specifically targets the ? chain of the CD8 differentiation antigen on mouse cells.

Primary Applications in In Vivo Studies

CD8+ T Cell DepletionThe 53-5.8 antibody is primarily used to deplete CD8+ T cells in living mice for experimental purposes. When administered in vivo, this antibody has been shown to completely deplete CD8+ T cells while notably preserving other cell populations. Importantly, it does not deplete CD8+ CD11c+ dendritic cells, making it a selective tool for targeting specific immune cell subsets.

Immunological ResearchResearchers utilize this antibody in studies investigating the role of conventional CD8??+ T cells in various disease models and immune responses. The depletion approach allows scientists to examine what happens when CD8+ T cells are removed from the immune system, helping to understand their contribution to immune responses, tumor immunity, and autoimmune conditions.

Mechanism and Specificity

The 53-5.8 clone recognizes an epitope on mouse CD8b.2 and is specific for Ly-3.2 phenotype, showing only low reactivity with Ly-3.1 strains such as AKR, C58, MRL, and PL mice. This specificity is important because CD8 can exist as either a heterodimer (CD8??) or homodimer (CD8??), and this antibody specifically targets the ? chain component.

Commercial Availability and Formats

The 53-5.8 clone is widely available from multiple suppliers in various formats optimized for in vivo use, including low endotoxin formulations specifically designed for animal studies. It's available in bulk quantities ranging from milligrams to multi-gram amounts, reflecting its common use in research. The antibody is also available in different conjugated forms for flow cytometry applications to monitor depletion efficiency.

This antibody has become a standard tool in immunological research, with over 100 product citations, demonstrating its widespread acceptance and reliability in the scientific community.

Commonly Used Antibodies and Proteins with 53-5.8

The 53-5.8 monoclonal antibody specifically targets the mouse CD8? chain (also known as Ly-3.2). Researchers often combine 53-5.8 with other antibodies and proteins to study CD8+ T cell biology, immune depletion, and cellular interactions. Here are some of the most commonly used reagents co-employed with 53-5.8 in the literature:

Anti-CD8? Antibodies

• 53-6.72: Frequently used alongside 53-5.8 to target the CD8? chain, enabling comprehensive coverage of the CD8 co-receptor (CD8?? heterodimer) on cytotoxic T cells and thymocytes.
• Other CD8? clones: Depending on the experimental context, additional anti-CD8? clones may be used, but 53-6.72 is most commonly paired with 53-5.8 for depletion or surface staining.

Additional T Cell Markers

• Anti-CD3: Used to pan-activate or identify T cells.
• Anti-CD4: For identifying and comparing CD4+ helper T cell populations.
• Anti-CD25: Used in functional studies, especially for regulatory T cell characterization.
• Anti-CD44, Anti-CD62L: For identifying memory and naive T cell subsets.

Depletion and Functional Studies

• Anti-CD11c: Sometimes used to distinguish CD8+ T cells from CD8+ CD11c+ dendritic cells, since 53-5.8 does not deplete dendritic cells.
• Anti-thymocyte markers: For studying T cell development in the thymus, as both CD8? and ? chains are expressed early.

Functional Assays and Recombinant Proteins

• MHC class I tetramers: To assess antigen-specific CD8+ T cell responses, often in combination with FACS staining using 53-5.8.
• Anti-IFN?, Anti-TNF?: For cytokine profiling of CD8+ T cell effector function.
• Anti-Granzyme B, Anti-Perforin: To evaluate cytotoxic activity.

Isotype Controls

• Rat IgG1: Used as an isotype control for 53-5.8 (which is a rat IgG1 antibody).
• Rat IgG2a: Isotype control for anti-CD8? (53-6.7, rat IgG2a).

Typical Experimental Combinations

A standard experimental setup might involve:

• Surface staining: 53-5.8 (CD8?), 53-6.72 (CD8?), CD3, CD4, CD44, CD62L for basic T cell subset analysis.
• Depletion experiments: 53-5.8 alone or with 53-6.72 for selective depletion of CD8+ T cells in vivo.
• Intracellular cytokine staining: 53-5.8 for CD8?, coupled with antibodies against cytokines such as IFN?, TNF?, Granzyme B.

Summary Table

Key Literature Context

• 53-5.8 is most often paired with 53-6.72 for comprehensive CD8+ T cell studies in mice, especially in depletion, functional assays, and multiparameter flow cytometry.
• When targeting CD8+ subsets, antibodies against activation markers (CD44, CD62L), cytokines (IFN?, TNF?), and cytotoxic molecules (Granzyme B) are frequently combined with 53-5.8 to profile T cell function.
• Dendritic cell distinction is important, as 53-5.8 does not deplete CD8+ CD11c+ dendritic cells.
• Isotype controls are essential, with rat IgG1 for 53-5.8 and rat IgG2a for 53-6.72.

These combinations reflect the standard toolkit for studying CD8+ T cell biology in the mouse system, leveraging 53-5.8 as a key reagent for identifying, depleting, or functionally analyzing CD8?-expressing cells.

Key findings from scientific literature citing clone 53-5.8, an anti-mouse CD8? monoclonal antibody, include its effectiveness for depleting CD8+ T cells in mice, the distinct effects it has on immune cell populations compared to anti-CD8? antibodies, and its utility in dissecting CD8+ T-cell–mediated immune functions and pathologies.

Essential findings and context from major citations:

• Specificity and Potency: Clone 53-5.8 targets the ?-chain (CD8?) of the mouse CD8 antigen, which is expressed predominantly on CD8+ T cells. Compared to anti-CD8? clones (e.g., 53-6.7), clone 53-5.8 more efficiently depletes CD8+ T cells in vivo and is recommended when maximal depletion is needed.

• Functional Implications of Depletion: Depletion using clone 53-5.8 leaves behind a population of CD8+ T cells with distinct functional and phenotypic properties. Surviving cells after anti-CD8? treatment may retain or even exhibit increased cytotoxic function, especially during primary immune responses. This has major implications for experiments analyzing the absence or function of CD8+ T cells as results can be confounded by the characteristics of the surviving population.

• Interpretation Challenges: The 53-5.8 antibody induces substantial CD8 internalization, complicating detection by flow cytometry post-depletion (staining intensity drops), and can make proper identification of residual CD8+ T cells difficult without careful gating strategies.

• Pathophysiological Applications: Clone 53-5.8 has been instrumental in demonstrating the pathogenic roles of CD8+ T cells in disease models. For instance, in a cerebral malaria mouse model, anti-CD8? (53-5.8) treatment depleted CD8+ T cells, reduced pulmonary vascular leakage, and improved survival, without affecting parasite burden—demonstrating the direct pathogenic role of CD8+ T cells in pulmonary complications.

• Comparisons and Limitations: Combining clone 53-5.8 with anti-CD8? mAbs does not increase depletion efficiency beyond that achieved with clone 53-5.8 alone, and using less effective clones (like anti-CD8? 53-6.7) can result in survival of memory-type T cells, potentially skewing long-term immunological studies.

Summary Table: Clone 53-5.8 vs. Anti-CD8? (53-6.7)

Additional Notes:

• Clone 53-5.8 is referenced as a gold standard tool for functional CD8+ T cell depletion in mouse immunology experiments.
• Researchers are advised to interpret results cautiously, considering possible residual and altered CD8+ populations after depletion.

In summary, the scientific literature characterizes clone 53-5.8 as a potent, widely used tool for mouse CD8+ T-cell depletion that profoundly influences subsequent immune responses and experimental outcomes.

Dosing regimens for clone 53-5.8 (anti-mouse CD8?) in mouse models typically range from 5 mg/kg administered weekly to 200–250??g per mouse, with variations depending on the experimental design and mouse strain.

• In syngeneic tumor models (e.g., MC38, CT26, B16-F10), clone 53-5.8 was administered at 5?mg/kg intraperitoneally on the day before, the day of implantation, and then weekly thereafter for CD8+ T cell depletion.
• Commercial antibody guides recommend using 200–250??g per mouse via intraperitoneal injection, typically 2–3 times per week for sustained depletion, especially in immune cell depletion studies.
• Clone 53-5.8 is described as completely depleting CD8+ T cells in vivo but not affecting certain dendritic cell subsets, supporting its specificity and utility across different immunological settings.

Key regimen variation factors:

• Tumor type and experimental schedule: Dosing frequency and start time may be modified depending on tumor model kinetics or the timing of immune manipulation (e.g., starting at day 4, 7, 14, or 25 post-implantation).
• Purpose of depletion: Some protocols use higher initial doses or more frequent dosing if rapid or thorough CD8+ T cell elimination is required (e.g., before transplantation, tumor challenge, or infection modeling).
• Mouse strain: Dose adjustments are sometimes made based on the weight or immune characteristics of specific mouse lines, but standard dosing regimens are widely applicable.

Summary Table—Regimen Examples:

Experimental protocols may tailor the dosing schedule to mouse strain, tumor type, or immune challenge timeline, but the above regimens represent standard practice in published studies and commercial guides.