Anti-Mouse PD-1 [Clone RMP1-30] — Purified in vivo GOLD™ Functional Grade

Clone RMP1-30 is a rat monoclonal antibody specific for mouse PD-1 (CD279), and its common in vivo applications in mice are primarily for detection and characterization of PD-1 expression, such as via flow cytometry and immunophenotyping, rather than for functional blocking of PD-1 signaling.

Key in vivo uses of RMP1-30 in mice include:

• Flow cytometric analysis of PD-1 expression on activated and unactivated T cells, B cells, dendritic cells, macrophages, and certain tumor cells.
• Immunophenotyping of immune cell populations in mice to monitor PD-1 surface protein, especially in studies on immune regulation and tumor immunology.
• Identification and isolation of PD-1+ cell populations (e.g., by FACS—fluorescence-activated cell sorting) for downstream analyses, such as transcript enrichment and functional studies.

Limitations and Important Notes:

• RMP1-30 is not considered suitable for in vivo functional PD-1 blockade (i.e., it does not block the PD-1/PD-L1 interaction in live animals). For functional/therapeutic in vivo blockade, other clones like 29F.1A12 are used preferentially.
• While RMP1-30 can bind PD-1 in in vivo settings, its primary role is as a detection and characterization tool rather than for manipulating PD-1-mediated immune pathways in therapeutic or mechanistic studies.

Summary Table: RMP1-30 in Vivo Applications in Mice

When planning experiments requiring functional modulation (blockade or agonism) of mouse PD-1 signaling in vivo, alternative clones should be selected.

RMP1-30 is commonly used in combination with several other antibodies and proteins in murine immune checkpoint research, particularly for validation, epitope mapping, and functional studies.

Anti-PD-1 Antibodies

29F.1A12 is one of the most frequently paired antibodies with RMP1-30. These two clones recognize overlapping populations, with co-staining studies showing that approximately 90-97% of PD-1-positive cells are dual positive for both antibodies. The 29F.1A12 antibody is particularly notable because it functions as a strong blocking antibody that can prevent PD-1/PD-L1 interactions, whereas RMP1-30 cannot block this interaction. This complementary functionality makes them useful together in experimental designs where one antibody blocks immune checkpoint signaling while the other serves as a detection reagent.

RMP1-14 is another anti-PD-1 antibody frequently used alongside RMP1-30. Like 29F.1A12, RMP1-14 has blocking capability, though it requires approximately 100-fold higher concentrations to achieve similar effects. RMP1-30 can be used to stain for PD-1 in experiments where RMP1-14 is employed therapeutically, though with an approximately 25% decrease in fluorescence when the therapeutic antibody is already bound to PD-1.

PD-L1 and Checkpoint Proteins

PD-L1 (also known as B7-H1) is frequently investigated in the same experimental systems as RMP1-30. As the primary ligand for PD-1, PD-L1 antibodies such as clone 10F.9G2 are often used in combination studies to understand the complete checkpoint interaction pathway. Additionally, PD-L2, the second ligand for PD-1, is relevant in these studies as it can also mediate inhibitory signals through PD-1 binding.

These antibody combinations are critical for dissecting immune checkpoint roles in health and disease, with researchers often using RMP1-30 for detection purposes while employing blocking antibodies like 29F.1A12 to modulate immune responses therapeutically.

Clone RMP1-30 is a monoclonal antibody against mouse PD-1 that has been widely cited in the scientific literature primarily as a PD-1 detection reagent in flow cytometry and immunophenotyping applications, but it does not function as a blocking antibody for the PD-1/PD-L1 interaction.

Key findings from scientific literature citations of RMP1-30:

• Specificity: RMP1-30 binds specifically to PD-1 on the surface of murine immune cells and tumor cells, with high specificity confirmed by lack of binding to PD-1 knockout cells.
• Epitope recognition and overlap: RMP1-30 and other anti-PD-1 clones (such as 29F.1A12 and RMP1-14) recognize overlapping but non-identical PD-1 epitopes. RMP1-30 can be used alongside 29F.1A12 and RMP1-14 for co-staining, with evidence for dual positivity in almost all PD-1^+^ cells.
• Blocking function: RMP1-30 does not block the interaction between PD-1 and PD-L1, and does not reverse PD-1 mediated inhibition of T cell signaling. This contrasts with clones 29F.1A12 and RMP1-14, which are capable of blocking activity. Studies confirmed that RMP1-30 does not enhance T cell activation via PD-1 pathway blockade.
• Use in combination with blocking antibodies: Because it is non-blocking, RMP1-30 is often used to stain or track PD-1 in the presence of blocking antibodies (e.g., 29F.1A12 or RMP1-14) to verify PD-1 status without interfering with the action of therapeutic or functional antibodies.
• Application in tumor and immune cell phenotyping: RMP1-30 is used to detect PD-1 on live and dead tumor cells, T cells, and other immune cell types in both 2D and 3D culture systems, revealing context-dependent differences in expression.
• Cooperative binding: There is some evidence of cooperative binding between RMP1-30 and certain other anti-PD-1 antibodies, potentially leading to increased fluorescence signal in antibody panels.

Additional notes:

• Agonist activity: Recent studies suggest potential for RMP1-30 (or closely related antibodies) to exhibit PD-1 agonist activity in specific in vitro or in vivo contexts, possibly modulating inflammation. However, evidence for robust agonist function in vivo remains limited and may depend on experimental conditions.
• Widespread citation: RMP1-30 is extensively cited as a key reagent in immuno-oncology studies, immune checkpoint validation, and comparative studies of anti–PD-1 antibodies in murine preclinical research.

In summary, RMP1-30 is a reliable, non-blocking anti–PD-1 clone for detection and phenotyping of murine PD-1, frequently used in combination antibody panels to dissect immune cell and tumor PD-1 expression, but is not used for functional PD-1 blockade in immunotherapy research.

Dosing regimens for clone RMP1-30 (anti-mouse PD-1 antibody) vary substantially across mouse models and research applications, with no single standardized protocol universally adopted. Most commonly, RMP1-30 is used for flow cytometry and immunophenotyping, not in vivo functional blockade; dosing and scheduling are therefore tied to experimental goals, cell numbers, and application type.

Key context for RMP1-30 dosing:

• Primary applications: Unlike clones such as 29F.1A12 or RMP1-14, RMP1-30 is typically utilized as a non-blocking antibody to detect PD-1 expression (e.g., by flow cytometry), rather than functional blockade or therapeutic intervention.
• Blocking activity: RMP1-30 does not block PD-L1 binding or stimulate T cell responses in vivo, so its dosing is usually not optimized for checkpoint blockade studies.

Dosing Regimen Variation

For flow cytometry:

• Typical working dilutions range from 1:100 to 1:200 of antibody stock (usually at 1 mg/ml).
• Commonly, 10 μl of dilution is used to stain 1 million cells in 100 μl buffer.

For in vivo use:

• Dosing guidelines are rarely published for RMP1-30, and when used, regimens are tailored to model demands (e.g., mouse strain, research question, marker expression).
• General recommendations for in vivo antibodies (other anti-PD-1 clones) are between 100–500 μg per mouse by intraperitoneal injection every 3–4 days, but these are specific to functional blocking clones and not directly applicable to RMP1-30.
• Leinco notes that RMP1-30 dosing "is not standardized and can vary considerably across different mouse models depending on the research application" and should be "optimized by the end user".

Essential Points

• No consensus dosing regimen exists for RMP1-30 in vivo; protocols are customized per experiment and are typically provided as starting points or require optimization by users.
• Published studies using RMP1-30 may report diverse protocols based on model type (i.e., tumor, autoimmune, or infectious disease), but generally do not specify standardized doses as they would for blocking antibodies.
• For detection or immunophenotyping, use recommended flow cytometry dilutions; for any functional or depletion study (rare for RMP1-30), start with comparable doses to other anti-PD-1 antibodies, but verify antibody activity and intent.

If your application is functional PD-1 blockade (e.g., immunotherapy or checkpoint studies), RMP1-30 is not appropriate due to its lack of blocking activity—use clones such as 29F.1A12 or RMP1-14 for those protocols.

In summary, doses of RMP1-30 are not standardized and vary with the mouse model and experimental objective, most commonly employed for PD-1 detection rather than in vivo modulation.