Anti-Mouse CD4 [Clone GK1.5] — Purified in vivo PLATINUM™ Functional Grade

The clone GK1.5 is a rat anti-mouse monoclonal antibody specifically targeting the CD4 antigen. It is commonly used for several in vivo applications in mice, including:

Common Applications

1. CD4+ T Cell Depletion: The primary use of GK1.5 is to deplete CD4+ T cells in vivo. This allows researchers to study the role of these cells in immune responses, disease progression, and therapeutic models. It is particularly useful in understanding immunity, disease, and therapeutic interventions.

2. Immune Response Studies: By depleting CD4+ T cells, researchers can examine how various diseases, such as autoimmune disorders and infections, progress without these cells. It helps in understanding T cell functions and their contributions to disease pathology.

3. Cancer Immunotherapy: Studies have shown that depleting CD4+ T cells, including regulatory T cells, can enhance antitumor immunity by increasing responses from effector T cells in mouse models of cancer.

4. Autoimmune Disease Models: GK1.5 has been used to reduce disease severity in models of autoimmune arthritis by eliminating CD4+ T cells, which play a significant role in triggering autoimmune responses.

5. Infectious Disease Models: It has been applied in models of HIV infection to reduce viral loads and disease duration, demonstrating its potential in treating infectious diseases.

Technical Applications

• Flow Cytometry: Used for identifying and quantifying CD4+ T cells.
• Immunohistochemistry: For staining and analyzing CD4+ T cell distribution in tissues.
• Immunoprecipitation: To isolate CD4+ T cells for further analysis.

These applications highlight the versatility and utility of the GK1.5 antibody in studying immune responses and diseases in mice.

When using GK1.5 (anti-mouse CD4 antibody) in research, scientists commonly pair it with antibodies or proteins targeting other immune cell markers to enable detailed immunophenotyping, cell depletion, or mechanistic studies. Frequently used antibodies or proteins in combination with GK1.5 include:

• CD8 antibodies (e.g., clone 53-6.7): These are used to identify or deplete cytotoxic T lymphocytes, often alongside GK1.5 to distinguish CD4+ versus CD8+ T cell populations during flow cytometry or depletion studies.
• CD3 antibodies (e.g., clone 145-2C11): CD3 marks all T cells and is routinely employed with CD4 and CD8 to comprehensively characterize T cell subsets.
• CD25 antibodies (e.g., clone PC61): CD25 targets the IL-2 receptor α-chain, useful for identifying regulatory T cells (CD4+ CD25+) in combination with GK1.5.
• Foxp3 antibodies: Foxp3 is used intracellularly to label regulatory T cells, when characterizing CD4+ Foxp3+ populations.
• Isotype control antibodies (e.g., rat IgG2b anti-KLH like LTF-2): Used to control for non-specific binding of GK1.5 or other rat monoclonals in flow cytometry or in vivo depletion.
• Other lineage markers (such as CD19 for B cells, CD11b/CD11c for myeloid cells): These enable broader immune cell profiling in multi-parameter flow cytometry.

A variety of GK1.5 competitors/blocking antibodies are also referenced in the literature, such as:

• RM4-5: Blocks GK1.5 binding to CD4+ T cells and is occasionally used to test binding specificity or blocking efficacy alongside GK1.5.
• RM4-4: Used for similar blocking studies but does not block GK1.5 binding.

In summary, the most commonly used antibodies or proteins with GK1.5 are:

• CD8 (cytotoxic T cell marker)
• CD3 (pan-T cell marker)
• CD25 and Foxp3 (regulatory T cell markers)
• Isotype controls (e.g., rat IgG2b anti-KLH)
• Competing CD4 antibodies (e.g., RM4-5, YTS 177, YTS 191)
• Other immune lineage markers (e.g., CD19, CD11b, CD11c)

These combinations allow effective immune monitoring, CD4+ T cell depletion studies, and immune subset discrimination in murine models.

Key Findings from Clone GK1.5 (Anti-mouse CD4) Citations

Depletion of CD4+ T Cells In Vivo

• Effective Depletion: Clone GK1.5 is widely used for the specific depletion of CD4+ T cells in experimental murine models. This application is instrumental for studying the functional contribution of CD4+ T cells to immune responses, autoimmune diseases, infection, and cancer immunotherapy.
• Disease Models: GK1.5-mediated CD4+ T cell depletion has been shown to reduce disease severity in models of autoimmune arthritis, demonstrating therapeutic potential by decreasing cytokine production and attenuating joint injury. In murine models of HIV, GK1.5 treatment reduced viral load and delayed disease progression. Combined with anti-CTLA-4, GK1.5 depletion also enhances antitumor immunity by targeting both CD4+ and regulatory T cells, thereby improving effector T cell responses.
• Functional Validation: GK1.5 is specifically formulated for in vivo use, ensuring high purity, ultra-low endotoxin levels, and absence of preservatives or carrier proteins, which minimizes confounding immune effects in animal studies.

Blockade of T Cell Activation and Function

• Receptor Blockade: The GK1.5 antibody competitively binds the CD4 co-receptor, inhibiting its interaction with MHC class II molecules on antigen-presenting cells, which is essential for optimal T cell activation. This can block CD4-mediated cell adhesion and helper T cell activation, both in vitro and in vivo.
• Competitive Binding: GK1.5 has been shown to compete with other anti-CD4 monoclonal antibodies (clones YTS 177 and YTS 191) for CD4 binding, confirming its specificity.
• Synaptic Block: In some contexts, such as fertilization, GK1.5 can also block CD4-dependent processes outside the immune system, such as cell adhesion during egg–sperm interaction.

Technical and Methodological Studies

• Detection and Staining: In addition to depletion, GK1.5 is used for flow cytometry, Western blotting, and immunohistochemistry to identify CD4+ T cells in various tissues and experimental conditions.
• Imaging Applications: Modified versions of GK1.5 (e.g., cys-diabody) have been explored for immunoPET imaging, allowing visualization of CD4+ T cell distribution in vivo with minimal perturbation to T cell biology.
• Control Considerations: Recommended isotype controls (e.g., rat IgG2b) and specialized buffers are provided for rigorous experimental design in in vivo studies.

Summary Table: Key Applications and Findings

Conclusion

Clone GK1.5 is a cornerstone tool in immunology research, primarily for depleting and blocking CD4+ T cell function in murine models. Its validated use spans basic research, therapeutic intervention studies, and advanced imaging, consistently demonstrating both specificity and reliability in delineating the roles of CD4+ T cells in health and disease.

Dosing Regimens of GK1.5 Across Mouse Models

Dosing regimens for the anti-mouse CD4 antibody clone GK1.5 vary significantly depending on the experimental objectives, mouse strain, route of administration, and desired immunological outcome. There is no universal "one-size-fits-all" protocol, and laboratories must optimize conditions for their specific model and research question.

Common Dosing Parameters

• Dose Range: The literature reports a wide range of doses, typically from 50 to 500 μg per injection, depending on the study. The most commonly cited regimens for in vivo CD4+ T cell depletion involve intraperitoneal injections of 200–250 μg per mouse.
• Frequency: Injections are often administered 2–3 times per week over the course of the experiment. However, some protocols may use a single injection for acute depletion or more frequent dosing for sustained effects.
• Route: Intraperitoneal injection is the standard route for systemic depletion studies. Other routes, such as intravenous, may be used in specialized protocols (e.g., for imaging), with much lower doses (e.g., 2–40 μg).
• Experimental Duration: The duration of treatment can range from a single administration to repeated dosing over several weeks, depending on whether the goal is acute or chronic depletion.

Factors Influencing Dosing Regimens

• Mouse Strain: Different strains may metabolize or respond to antibodies differently, affecting both efficacy and potential side effects.
• Purpose of Study: For example, studies aiming for complete depletion (e.g., in autoimmune disease models) may use higher or more frequent doses than those using GK1.5 for functional blockade or partial suppression of CD4+ T cells.
• Age and Health Status: Younger or immunocompromised mice may require adjusted dosing to avoid excessive toxicity.
• Off-Target Effects: GK1.5 can also bind to non-T cell populations expressing low levels of CD4, which may necessitate lower or more carefully titrated doses to minimize unintended depletion events.

Example Regimens

Optimization and Validation

It is essential for researchers to perform dose-response experiments to determine the optimal regimen for their specific experimental setup, taking into account mouse strain, age, health, and the primary research objective. Validation by flow cytometry or histological assessment of depletion efficiency is recommended to confirm the adequacy of the chosen regimen.

Conclusion

GK1.5 dosing regimens in mice are highly context-dependent, with typical doses ranging from 50 to 500 μg, most commonly 200–250 μg intraperitoneally 2–3 times per week for systemic CD4+ T cell depletion. However, the optimal protocol should always be determined empirically for each laboratory’s specific model and goals.