Anti-Mouse NKG2A/C/E – Purified in vivo GOLD™ Functional Grade

Clone 20D5 is widely used in vivo in mice to study and manipulate the function of natural killer (NK) cells and CD8+ T cells by targeting the NKG2A, NKG2C, and NKG2E receptors, primarily within the context of cancer immunology and immune regulation.

Key in vivo applications include:

• Immune checkpoint inhibition in cancer models: The 20D5 antibody blocks the inhibitory NKG2A receptor, which can unleash both NK cell and CD8+ T-cell activity, thereby promoting anti-tumor immunity. This makes it a valuable tool for preclinical studies of cancer immunotherapy and immune checkpoint blockade.
• Functional depletion or modulation of NK and NKT cells: By binding to NKG2A/C/E on NK and NKT cells, 20D5 can be used to modulate or study these cell populations in vivo, either through blockade or potential depletion (depending on the isotype and experimental setup).
• Synergy with adoptive cell therapy: In some studies, 20D5 has been used in combination with intratumoral injection of activated NK cells to achieve synergistic anti-tumor effects in solid tumor models.
• General characterization of NKG2A/C/E-expressing cell subsets: 20D5 is also employed to phenotype or track NKG2A/C/E-positive NK, NKT, and some CD8+ T cells in vivo, aiding studies on their role in infection, autoimmunity, or tumor surveillance.

Summary table of applications:

Important considerations:

• DBA/2J mice lack CD94 and do not express CD94/NKG2 receptors, so clone 20D5 is ineffective in this strain.
• 20D5 is generally non-depleting in vivo, acting primarily as a blocking antibody, but this can depend on the Fc region of the antibody preparation.
• Always verify dosing and compatibility with mouse strains and experimental goals.

In summary, clone 20D5 is primarily used for checkpoint blockade, functional modulation, and characterization of NKG2A/C/E-positive lymphocytes in a variety of in vivo mouse models, especially in cancer and immunotherapy research.

Commonly used antibodies or proteins in combination with 20D5 (anti-mouse NKG2A/C/E) in the literature include anti-PD-1, anti-PD-L1, anti-asialo-GM1, and anti-CD8 antibodies. These combinations are primarily employed to study immune modulation and tumor immunity.

Key details:

• Anti-PD-1 and anti-PD-L1: Frequently paired with 20D5 to investigate interactions between NK cell inhibitory checkpoints (NKG2A/C/E) and PD-1/PD-L1 pathways, particularly in tumor immunology models.
• Anti-asialo-GM1: Commonly used for NK cell depletion, enabling assessment of NK cell-specific responses relative to NKG2A/C/E blockade.
• Anti-CD8: Utilized to discriminate between effects on CD8+ T cells and NK cells in functional assays involving NKG2A/C/E signaling.
• Additional combinations: Studies frequently examine the synergistic effects of co-injecting NKG2A-neutralizing antibodies (such as 20D5) with activated NK cells for enhanced anti-tumor activity.

These antibodies are widely chosen because they target pathways or cell types that interact with NKG2A/C/E-expressing cells, allowing precise dissection of immune cell roles in cancer and infection models.

Clone 20D5 is a rat monoclonal antibody widely used in immunology research to detect and functionally block mouse NKG2A, NKG2C, and NKG2E receptors, which are members of the C-type lectin-like receptor family expressed primarily on NK and NKT cells. Key findings from scientific literature using 20D5 can be summarized as follows:

• Specificity and Applications:
  Clone 20D5 binds to the mouse NKG2A, NKG2C, and NKG2E isoforms and is used to identify and study NK and NKT cell subsets across most mouse strains except DBA/2J, which lack the required CD94 co-receptor. This antibody is frequently employed in flow cytometry, functional assays, and in vivo studies to both deplete and analyze NK cell populations.

• Functional Blockade and Immunotherapy:
  Blocking NKG2A signaling with clone 20D5 enhances NK and CD8+ T cell anti-tumor responses in mouse tumor models by releasing inhibitory brakes on these cells. This has provided preclinical basis for targeting NKG2A as a novel immune checkpoint inhibitor. Anti-NKG2A/C/E therapy with 20D5 has been shown to promote anti-tumor immunity and is of significant interest in cancer immunotherapy research.

• Checkpoint in Viral Immunity and Exhaustion:
  Studies show that 20D5 is valuable in models of chronic viral infection, such as hepatitis C virus (HCV), where it is used to explore the role of NKG2A as an exhaustion checkpoint on NK cells. NKG2A+ NK cells are implicated in contributing to viral persistence and immune suppression.

• Mechanistic Insights:
  Research using 20D5 established that NKG2A typically functions as an inhibitory receptor that recognizes the nonclassical MHC class I molecule Qa-1 presenting the Qdm peptide, whereas NKG2C and NKG2E can transduce activating signals. Clone 20D5 has been instrumental in mapping these signaling pathways and confirming cell-surface protein expression in various genetic backgrounds.

• Synergy with Other Immunotherapies:
  Combination blockade of NKG2A (using 20D5) with anti–PD-1 or PD-L1 antibodies can further enhance anti-tumor efficacy, attributed to cooperative disinhibition of cytotoxic lymphocytes. These findings support ongoing research into combinatorial checkpoint inhibition strategies for cancer treatment.

• Technical Limitations:
  Because clone 20D5 reacts with all three isoforms (NKG2A/C/E), its use requires careful interpretation in experiments where distinguishing between these isoforms is critical. In some studies, the effects of 20D5 are attributed specifically to NKG2A when the other isoforms are not functionally expressed or relevant.

In summary, clone 20D5 is foundational for studying NK cell biology, checkpoint inhibition, and immune regulation in murine models, especially in the contexts of cancer and viral immunity. It remains an essential tool for both basic and translational immunological research.

Dosing regimens for clone 20D5 (anti-mouse NKG2A/C/E) antibodies in mouse models vary based on the experimental design, mouse strain, and presence of disease (e.g., tumor vs. normal). Standard regimens typically involve intraperitoneal (i.p.) injections with single or repeated doses ranging from 200–300 μg per mouse.

Key details on dosing regimens across different mouse models:

• Common Dose: Most studies use 200–300 μg per mouse per injection, administered via intraperitoneal route.
• Typical Schedule: Doses can be given as a single bolus or as part of a repeated schedule; for example, every 3 days or on specific study days post-tumor inoculation.
• Tumor Models: In B16F10 melanoma tumor-bearing mice, 200 μg of 20D5 was administered on days 7, 10, 13, and 16 post-irradiation/tumor challenge.
• Other Models: In studies on T cell and NK cell activation using Con A or α-GalCer challenge, a single 300 μg i.p. dose was given 2 days before the challenge.
• Genetic Background Considerations: Clone 20D5 reacts with NKG2A/C/E antigens in most mouse strains except DBA/2J, which lack CD94 and do not express CD94/NKG2 receptors.
• Route of Administration: While i.p. injection is most common, intratumoral injection may be used for local effects in solid tumor models.

Summary Table: Example 20D5 Regimens in Mouse Models

Additional relevant details

• Mouse strain selection impacts usefulness: DBA/2J mice not suitable due to lack of CD94/NKG2 expression.
• Antibody source and formulation: Most commercially available 20D5 antibodies are functional grade and certified for low endotoxin, suitable for in vivo studies.
• Adjustment for study context: Total number of injections, interval, and combination with other therapies (e.g., PD-1 blockade) should align with research aims.

In summary:

• The most widely used dosing regimens for clone 20D5 in mice are 200–300 μg/injection i.p., either as single or repeated injections.
• Frequency and timing depend on the experimental model (tumor, immune activation, co-therapy), and most protocols use intervals of several days between doses.
• Always verify strain compatibility, as not all mice express the relevant antigenic complex required for clone 20D5 binding.