Anti-Mouse PD-L1 [Clone 10F.9G2] — Purified in vivo PLATINUM™ Functional Grade

In Vivo Use of Clone 10F.9G2 in Mouse Studies

Clone 10F.9G2 is a rat monoclonal antibody that specifically targets mouse PD-L1 (programmed death ligand 1, also known as B7-H1 or CD274). This antibody is widely used in vivo for its ability to detect, modulate, or block PD-L1-mediated immune responses in mice.

Key Applications

• Blocking PD-L1 Immune Checkpoint: 10F.9G2 is commonly used to block the interaction between PD-L1 and its receptor PD-1, which is critical for understanding immune checkpoint biology in vivo. By inhibiting this pathway, the antibody disrupts immunosuppressive signals, allowing for enhanced T cell activity and increased immune response against certain targets, including tumors or autoantigens.
• Inflammation and Autoimmunity Models: In studies of autoimmune diabetes, administration of 10F.9G2 (or the related clone 10F.2H11) to mice accelerated disease onset and increased the incidence and severity of insulitis when CD8+ T cells were transferred, demonstrating its role in removing PD-L1-mediated immune inhibition and promoting effector T cell function in vivo.
• Cancer Immunotherapy Research: In mouse models of melanoma, 10F.9G2 treatment can transiently inhibit tumor growth by blocking PD-1/PD-L1 interaction, thereby preventing tumor immune evasion and promoting tumor rejection by the adaptive immune system.
• Blocking Alternative PD-L1 Pathways: Notably, 10F.9G2 also blocks the PD-L1:B7-1 (CD80) interaction, which is distinct from the PD-1 pathway, adding to its versatility in dissecting immune regulation mechanisms in mice.

Methodological Notes

• Administration: The antibody is typically administered intraperitoneally or intravenously to mice at dosages and schedules tailored to the experimental design.
• Specificity: 10F.9G2 is highly specific to mouse PD-L1, making it a key tool for studies in murine systems, but not directly translatable to human studies without further engineering.
• Functional Outcomes: The in vivo effects of 10F.9G2 include increased T cell infiltration (e.g., into pancreatic islets in diabetes models), enhanced cytokine production, and, in tumor models, delayed tumor progression via immune-mediated mechanisms.
• Comparative Studies: 10F.9G2 is often compared with other anti-PD-L1 clones (e.g., MIH6) to assess differences in binding, blocking efficiency, and biological outcomes in vivo.

Summary Table: Key In Vivo Uses of 10F.9G2

Conclusion

Clone 10F.9G2 is a foundational tool for in vivo mouse studies of immune checkpoint biology, inflammation, autoimmunity, and cancer immunotherapy. Its ability to block both PD-L1:PD-1 and PD-L1:B7-1 interactions makes it uniquely valuable for dissecting the roles of PD-L1 in immune regulation and disease progression in murine models.

The 10F.9G2 antibody is commonly used alongside several other antibodies and proteins in research studies, particularly in immunology and cancer research contexts.

Related Anti-PD-L1 Antibodies

10F.2H11 is the most frequently co-mentioned antibody with 10F.9G2 in the literature. These two antibodies have distinct functional properties despite both targeting PD-L1. While 10F.9G2 acts as a "dual-blocker" that prevents binding of PD-L1 to both PD-1 and B7-1 receptors, 10F.2H11 specifically blocks only the PD-L1:B7-1 interaction while leaving PD-L1:PD-1 binding intact. Both antibodies have comparable affinities for PD-L1 and show similar staining patterns on PD-L1-transfected cells.

Target Proteins and Receptors

The primary protein targets associated with 10F.9G2 studies include:

PD-1 (Programmed Death-1) - This is one of the key receptors that PD-L1 binds to, and 10F.9G2's ability to block this interaction is central to its functional effects.

B7-1 (CD80) - Another important binding partner of PD-L1. The dual-blocking capacity of 10F.9G2 to disrupt both PD-L1:PD-1 and PD-L1:B7-1 interactions makes it particularly valuable for functional studies.

CD274 - This is the mouse gene that encodes PD-L1, and 10F.9G2 is specifically designed to recognize this protein.

Comparative Research Applications

In diabetes research, both 10F.9G2 and 10F.2H11 have been used together to understand different aspects of PD-L1 signaling pathways. Studies have shown that these antibodies have distinct effects when used to treat NOD mice, with 10F.9G2 showing more potent effects in accelerating diabetes progression compared to 10F.2H11.

Technical Considerations

29F.1A12 is mentioned as a monoclonal antibody specific for mouse PD-1, often used in comparative studies alongside 10F.9G2 to examine both sides of the PD-1/PD-L1 interaction pathway.

The literature frequently references isotype controls such as rat IgG2b when using 10F.9G2, as this antibody was originally developed as a rat monoclonal antibody, though recombinant mouse versions are now available.

Key Findings from Scientific Literature on Clone 10F.9G2

Epitope Specificity and Functional Blockade

• Epitope Recognition: Clone 10F.9G2 targets PD-L1 (CD274) and recognizes a distinct epitope from another well-characterized anti-PD-L1 antibody, 10F.2H11. Competitive binding assays show that 10F.9G2 does not block 10F.2H11 binding, and vice versa, indicating non-overlapping epitopes on PD-L1.
• Dual Functional Blockade: Unlike 10F.2H11, which only blocks PD-L1:B7-1 interactions, 10F.9G2 is a “dual blocker”—it inhibits both PD-L1:PD-1 and PD-L1:B7-1 interactions. This was demonstrated using in vitro adhesion assays with cells expressing PD-L1 and wells coated with PD-1 or B7-1 fusion proteins.
• Overlap with Other Dual Blockers: 10F.9G2’s binding site overlaps with another dual-blocking antibody, MIH5, as pre-incubation with 10F.9G2 significantly reduces MIH5 binding, while 10F.2H11 does not.

In Vivo and Therapeutic Effects

• Tumor Growth Inhibition: In syngeneic mouse tumor models (e.g., D2F2, 4T1, CT26), treatment with mAb 10F.9G2 significantly inhibits tumor growth compared to PBS controls, with comparable efficacy to certain PD-L1 peptide vaccines in the 4T1 model. Specifically, both 10F.9G2 and peptide vaccines prolonged survival and reduced tumor volume, but in some models (e.g., D2F2), peptide vaccines showed superior tumor inhibition compared to 10F.9G2.
• Survival Benefit: Mice treated with 10F.9G2 showed prolonged survival rates across multiple tumor models, indicating a robust anti-tumor immune response.
• Comparative Efficacy: While 10F.9G2 is effective, certain peptide vaccines (e.g., TT3-PD-L1(130)) induced higher tumor growth inhibition (%TGI) and were more effective in some models than 10F.9G2, particularly in the D2F2 and CT26 models.

Technical and Immunological Properties

• Antibody Type and Specificity: 10F.9G2 is a rat monoclonal antibody specific to mouse PD-L1 (CD274), validated for applications such as flow cytometry.
• Affinity and Staining: 10F.9G2 and 10F.2H11 have comparable affinities for PD-L1 and similar staining intensity on PD-L1-transfected cells, but their functional blockade differs due to epitope specificity.

Summary Table

Conclusion

Clone 10F.9G2 is a rat monoclonal antibody that uniquely blocks both PD-L1:PD-1 and PD-L1:B7-1 interactions due to its specific epitope recognition, distinguishing it from other anti-PD-L1 antibodies like 10F.2H11. It demonstrates significant anti-tumor activity in mouse models, though some PD-L1-targeted vaccines can outperform it in certain contexts. Its properties make it a valuable tool for studying PD-L1 function and immunotherapy mechanisms.

Dosing regimens for clone 10F.9G2 (anti-mouse PD-L1 antibody) typically range between mouse models, but share core parameters informed by immunotherapy research. Most studies and dosing guides recommend the following protocol:

• Dose per mouse: 100–250??g per injection.
• Route: Intraperitoneal (i.p.) injection.
• Frequency: 2–3 times per week; some protocols, including tumor studies, use daily dosing for up to 14 days at 250??g/dose.

Common applications:

• Used primarily for cancer immunotherapy in syngeneic tumor models (e.g., MC38, B16 melanoma).
• Occasionally applied in infection models for T cell enhancement.

Pharmacokinetic variations and mouse strain differences:

• In CD1 outbred mice, pharmacokinetic studies used 1?mg/kg and 10?mg/kg twice weekly, observing substantial inter-dose drops in serum level between the fourth and sixth dose, likely due to anti-drug antibody (ADA) formation, which can significantly affect exposure and elimination.
• Enhanced elimination after repeated dosing may be species- or strain-specific and can relate to immunogenicity (rat variable sequence in the antibody triggers mouse ADA responses).

Specific model regimens:

• Syngeneic tumor models: 200–250 ?g per dose, given 2–3 times per week (or daily for up to 2 weeks in intensive protocols).
• CD1 pharmacokinetic studies: 10?mg/kg, twice weekly dosing; lower exposures due to rapid elimination relative to similar clones.

Considerations:

• ADA formation is a significant variable—some mouse strains or repeated dosing cycles may clear the antibody faster, necessitating dose or schedule adjustment.
• The chosen dose and schedule may also be tailored for the aggressiveness of the tumor, the sensitivity of the mouse strain to immunogenic responses, and the desired immune activation profile.

In summary:
Most studies and guides recommend 100–250??g per mouse, i.p., 2–3 times per week, but dosing in some models may escalate to daily injections at the upper end of the range, or employ weight-based dosing (e.g., 10 mg/kg) to match specific pharmacokinetic or immunogenicity requirements. Dose customization across mouse models reflects differences in strain, tumor burden, and duration of study.