Anti-Mouse CD279 (PD-1) [Clone RMP1-14] — Purified in vivo PLATINUM™ Functional Grade

Use of RMP1-14 in In Vivo Mouse Studies

RMP1-14 is a highly characterized rat IgG2a monoclonal antibody developed for blocking mouse PD-1 (Programmed Cell Death Protein 1), an immune checkpoint receptor critical for regulating T cell activation and function. This clone is one of the most widely used and established tools for in vivo research in mice, particularly in the fields of cancer immunotherapy and immune regulation.

Application in In Vivo Models

• Cancer Immunotherapy: RMP1-14 is commonly administered in syngeneic tumor models (e.g., MC38 colon carcinoma or B16 melanoma) to evaluate the efficacy of PD-1 blockade as a therapeutic strategy. The antibody effectively disrupts the PD-1/PD-L1 (and PD-L2) interaction, resulting in enhanced T cell activation and anti-tumor immune responses.
• Dosing and Administration: Typical administration is intraperitoneal, with doses commonly ranging from 200–500 ?g per injection, delivered every 3–4 days. The optimal dose and frequency may need adjustment depending on the tumor model, mouse strain, and experimental objectives.
• Mechanistic Studies: Besides therapeutic outcomes, RMP1-14 is used to dissect mechanistic aspects of PD-1 signaling in vivo, including its role in immune tolerance and autoimmunity.

Experimental Considerations

• Animal Model Selection: The choice of mouse strain and tumor model is crucial, as response to PD-1 blockade can vary significantly between models, strains, and environmental conditions.
• Monitoring and Adverse Effects: PD-1 blockade, including with RMP1-14, can induce immune-related adverse events, particularly autoimmunity or toxicity. Continuous monitoring of animal health and adjustment of dosing are recommended to manage these risks.
• Limitations: RMP1-14 is specific for in vivo blocking of PD-1/PD-L1/PD-L2 interactions in mice and is not typically used for in vitro assays, immunohistochemistry, or flow cytometry. Other clones (such as 29F.1A12 or J43) are preferred for those applications.

Key Properties

• Host: Rat
• Isotype: IgG2a kappa
• Specificity: Recognizes an epitope on mouse PD-1, expressed by activated T, B cells, and some myeloid cells.
• Endotoxin Level: Ultra-low endotoxin preparations are available, which is important for in vivo studies to minimize inflammation.

Comparison with Other Clones

Summary

RMP1-14 is a trusted tool for in vivo PD-1 blockade studies in mice, primarily for cancer immunotherapy research. Its use is characterized by robust, reproducible blocking of PD-1/PD-L1 interaction, leading to enhanced anti-tumor immunity, but it requires careful model selection, dosing, and health monitoring due to potential immune-related side effects. It is not suited for in vitro or diagnostic applications, which are better served by alternative clones.

The correct storage temperatures for the sterile packaged clone RMP1-14 (anti-mouse PD-1 antibody) are as follows:

• Short-term storage: Store at 4°C for up to 1-2 weeks.
• Long-term storage: Store at -20°C for up to 12 months or at -70°C for longer term storage.

It is crucial to avoid repeated freeze-thaw cycles to maintain the integrity of the antibody.

In the literature, RMP1-14 is often used alongside other antibodies and proteins to study immune checkpoints, particularly PD-1 and PD-L1 interactions. Here are some commonly used antibodies and proteins used in conjunction with RMP1-14:

1. 10F.9G2 (Anti-mouse PD-L1): This antibody is often used to block PD-L1, the ligand for PD-1, and is effective in combination with anti-PD-1 antibodies like RMP1-14 for studying immune checkpoint blockade.

2. 29F.1A12 (Anti-mouse PD-1): Known for its high affinity and potent blocking ability, 29F.1A12 is used in studies to compare or combine with RMP1-14, particularly in models where strong PD-1 blockade is desired.

3. RMP1-30 (Anti-mouse PD-1): While RMP1-30 does not block PD-1/PD-L1 interaction effectively, it can be used for staining PD-1, especially in experiments involving RMP1-14.

4. J43 (Anti-mouse PD-1): This clone is sometimes compared to RMP1-14 in studies to assess efficacy in preclinical tumor models.

These antibodies are used to explore the mechanisms of PD-1/PD-L1 checkpoint blockade and to evaluate their potential in cancer immunotherapy and other immunological studies.

Key Findings on Clone RMP1-14 in Scientific Literature

Clone RMP1-14 is a rat IgG2a monoclonal antibody with high specificity and affinity for mouse programmed death-1 (PD-1), a critical immune checkpoint receptor. Its scientific utility is grounded in a robust preclinical record, particularly in cancer immunotherapy research.

Mechanism and Functional Properties

• Effective PD-1 Blockade: RMP1-14 potently blocks the interaction between PD-1 and its ligands, PD-L1 and PD-L2, thereby preventing the inhibitory signaling that suppresses T cell activation.
• Restoration of T Cell Function: By blocking the PD-1 pathway, RMP1-14 restores T cell proliferation and cytokine production, leading to enhanced anti-tumor immune responses in murine models.
• High Affinity and Specificity: RMP1-14 was selected for its strong binding to mouse PD-1 and minimal off-target effects, making it a reliable tool for in vivo studies focused on immune checkpoint modulation.

Preclinical Efficacy in Tumor Models

• Broad Anti-Tumor Activity: RMP1-14 has demonstrated significant tumor suppression across diverse cancer types, including glioma (65% suppression), melanoma (up to 80% when combined with photodynamic therapy), breast cancer (75% with NOS inhibitor), and mesothelioma (82% with combination therapy).
• Enhanced Combination Therapy Outcomes: In combination with modalities like photodynamic therapy, RMP1-14 has shown synergistic effects, leading to greater tumor control and survival benefits in animal models.
• Comparative Advantage: RMP1-14 outperforms other mouse-specific anti-PD-1 clones (e.g., J43) in terms of tumor growth inhibition and survival in some models, attributed to its higher binding affinity and stronger effector functions. However, it is outperformed by cross-reactive human/mouse clones (e.g., EH12.2H7) in humanized models, highlighting the importance of species specificity in experimental design.
• Consistent and Validated Protocols: The clone’s dosing and administration protocols are well established across tumor models, reducing the need for optimization and improving reproducibility in preclinical studies.

Applications in Research and Diagnostics

• Tool for Immune Microenvironment Studies: RMP1-14 is widely used to dissect the role of PD-1 in the tumor microenvironment, enabling researchers to model human-like immune responses in mice.
• Biomarker Discovery: It has facilitated the identification of immune-resistant tumor variants and supports the development of diagnostic tools for patient stratification in PD-1-targeted therapies.
• Enabling Translational Research: While primarily specific to mouse PD-1, RMP1-14’s extensive preclinical data provide a foundation for translational studies, though cross-reactive clones may be needed for humanized models.

Limitations

• Species Specificity: RMP1-14 is specific to mouse PD-1 and does not cross-react with human PD-1, limiting its direct translational applicability to human systems.
• Comparative Considerations: The choice between RMP1-14 and other clones should be guided by the research question, experimental model, and desired endpoints, with careful attention to binding properties and species reactivity.

Summary Table: Select Tumor Suppression Data with RMP1-14

Conclusion

Clone RMP1-14 is a cornerstone of preclinical cancer immunotherapy research, valued for its specificity, efficacy, and well-characterized use in murine models. Its ability to restore anti-tumor immunity and synergize with other therapies has made it a gold standard for exploring PD-1 blockade, though researchers must consider species specificity and clone selection based on their experimental goals.