Anti-Mouse CD16.2 (Clone 9E9) – Purified in vivo PLATINUM™ Functional Grade

Clone 9E9 is primarily used in vivo in mice to functionally block the FcγRIV (CD16.2) receptor, and, under certain conditions, also blocks FcγRIII, thereby allowing researchers to dissect the roles of these receptors in immune effector functions—especially those mediated by IgG2a and IgG2b antibodies.

Key in vivo applications include:

• Investigating the role of FcγRIV in IgG2a/IgG2b-mediated responses, such as platelet clearance and processes like B cell depletion, antibody-dependent cellular cytotoxicity (ADCC), and immune complex clearance.
• Studying the contribution of FcγRIV and FcγRIII to immune effector and inflammatory responses by selectively blocking these receptors. Clone 9E9 is unique because it blocks FcγRIII only when it also binds FcγRIV on the same cell, thus allowing for concurrent inhibition of both, which is particularly relevant for studies on neutrophil and macrophage activation in vivo.
• Analyzing myeloid cell functions (monocytes, macrophages, neutrophils) in various disease models, including:
  • Autoimmune models, such as nephrotoxic nephritis, to determine the role of FcγRIV-mediated activation in pathology.
  • Cancer immunotherapy, to study how myeloid cells and FcγR engagement affect clearance of tumor cells during monoclonal antibody treatment (e.g., depletion of antibody-tagged B cells or tumors following mAb therapy).
• Blocking experiments to dissect receptor redundancy and cooperation in Fc receptor signaling pathways, often used alongside genetic knockout models to parse out the distinct and overlapping functions of mouse Fcγ receptors.

Other relevant details:

• FcγRIV is expressed on monocytes, macrophages, dendritic cells, neutrophils, and other myeloid cells but is absent from lymphoid populations.
• 9E9-mediated blockade is critical for defining which FcγRs mediate responses to IgG2a/IgG2b-based therapeutics (such as antibody-mediated clearance or depletion).

In summary, clone 9E9 is widely used in vivo in mice to precisely inhibit FcγRIV-dependent functions and, in specific contexts, FcγRIII, making it a valuable tool for immunological research in autoimmunity, oncology, and inflammation.

Commonly Used Antibodies and Proteins with 9E9 in the Literature

9E9 refers to a clone of monoclonal antibody primarily targeting mouse CD16.2 (FcγRIV), used for blocking, flow cytometry, and functional studies in immunology research. In the literature, 9E9 is often used in tandem with other antibodies targeting different Fc receptors or cell markers to dissect immune complex-mediated responses, cell type-specific roles, and in vivo mechanisms.

Frequently Co-Used Antibodies

• Anti-FcγRIII Antibodies (e.g., 2.4G2): Used to block FcγRIII (CD16/CD32) to isolate the role of FcγRIV in immune responses. 2.4G2 is a rat IgG2b antibody that binds both FcγRIIb and FcγRIII and is often used as a blocking reagent in combination with 9E9 and other cell surface markers.
• Anti-FcγRIIB Antibodies (e.g., K9.361): Used to specifically target the inhibitory FcγRIIB receptor. For example, one study used clone K9.361 (Ly17.2-specific) in combination with 9E9 and other Fc receptor antibodies to measure receptor expression on myeloid cells.
• Anti-FcγRI (CD64) Antibodies (e.g., X54-5/7.1.1): Mouse IgG1 antibodies detecting FcγRI are used alongside 9E9 for comprehensive profiling of Fc receptor expression on immune cells.
• Anti-MXRA8 Antibodies: A different clone 9E9 also exists for MXRA8 (a completely unrelated protein), but this is not typically used in the context of Fc receptor biology.

Frequently Co-Used Proteins and Markers

• Mouse Immunoglobulins (e.g., IgG2a, IgG2b): Used in functional assays to study immune complex (IC) binding and effector cell activation, especially since FcγRIV (targeted by 9E9) is critical for IgG2a- and IgG2b-mediated responses.
• Cell Surface Markers (e.g., B220, CD11b, F4/80): Used for cell-specific phenotyping. For instance, studies often stain for F4/80 (macrophages) and CD11b (myeloid cells) together with Fc receptor antibodies to identify receptor expression on specific cell types.
• Complement Components: Some studies investigate the interplay between complement activation and Fc receptor-mediated effector functions, though complement proteins themselves are less commonly co-stained directly with 9E9.
• Isotype Controls (e.g., Armenian Hamster IgG): Used as negative controls to confirm specificity in flow cytometry and blocking experiments.

Example Table: Antibodies/Proteins Frequently Used with 9E9

Summary

In the literature, 9E9 is most commonly paired with antibodies against other Fc receptors (2.4G2 for FcγRIIb/III, K9.361 for FcγRIIB, X54-5/7.1.1 for FcγRI) to dissect the roles of individual FcγRs in immune responses. It is also used alongside general myeloid and lymphoid markers (B220, F4/80, CD11b) for phenotypic characterization. Complement proteins and immune complexes (especially mouse IgG2a/IgG2b) are used in functional assays to probe FcγRIV-dependent effector mechanisms in vivo and in vitro.

Key Findings from Clone 9E9 Citations in Scientific Literature

Anti-mouse FcγRIV clone 9E9 is a critical tool for dissecting the role of FcγRIV (CD16.2) in immune responses, especially in the context of IgG2a/c-mediated effector functions. The following are the main findings from peer-reviewed studies and technical summaries that cite the use of clone 9E9:

• Specific Blockade of FcγRIV: Clone 9E9 is widely used to specifically inhibit FcγRIV function in mouse models, allowing researchers to study its role in antibody-mediated immune responses independent of other Fc receptors.
• Reduction in Immune Pathology: Administration of anti-FcγRIV (clone 9E9) significantly decreases FcγRIV expression on splenic antigen-presenting cells (APCs), thereby reducing IgG2c-mediated enhancement of red blood cell (RBC) alloantibody production to background levels. This blockade can ameliorate pathology in models of autoimmune hemolytic anemia, rheumatoid arthritis, and nephritis, suggesting FcγRIV as a potential therapeutic target.
• Selective Effects on Fc Receptor Expression: While clone 9E9 primarily targets FcγRIV, it also reduces surface expression of FcγRIII and FcγRII (but not FcγRI), complicating attribution of effects solely to FcγRIV blockade. However, genetic studies with FcγRIII-deficient mice indicate that FcγRIV is the dominant mediator of the IgG2c-mediated enhancement of alloimmunization.
• Mechanistic Insights: Ligation of FcγRIV by IgG2a/c immune complexes promotes cellular activation (e.g., upregulation of CD86) and enhances antigen presentation, leading to increased T cell activation and proliferation.
• Technical Utility in Flow Cytometry: Clone 9E9 is used to block FcγRIV specifically when studying other Fc receptors by flow cytometry, minimizing false positives and ensuring unambiguous receptor quantification.
• In Vivo Functional Studies: The antibody is available in functional grades suitable for in vivo administration, supporting its use in animal models to probe FcγRIV’s biological functions.

Summary Table: Key Effects of Clone 9E9

Conclusion

Clone 9E9 is the gold-standard tool for selectively inhibiting FcγRIV in mice, enabling clear demonstration of its critical role in IgG2a/c-mediated immune responses, autoimmune pathology, and antigen presentation. While there are some off-target effects on FcγRIII and FcγRII expression, genetic evidence supports the centrality of FcγRIV in these pathways. These findings make FcγRIV (and by extension, clone 9E9) highly relevant for both basic immunology and therapeutic development.

There is currently no published, peer-reviewed data in the search results that directly describes how the dosing regimen of clone 9E9 (anti-mouse CD16.2/FcgRIV) varies across different mouse models. The available information provides details on the specificity and applications of clone 9E9, such as its reactivity with mouse macrophages, neutrophils, and dendritic cells, but does not specify dosing regimens for in vivo use in any mouse model.

A vendor listing for clone 9E9 also states that there is no substantial evidence of major differences in dosing regimens between mouse strains, but cautions that details could vary based on experimental context. This suggests that while there is no well-documented variation, it remains possible that minor adjustments might be needed depending on the specific model or experimental design. No standard or suggested dose for clone 9E9 in any mouse model is provided in the available literature.

Expert Guidance

• No established dosing guidelines: Unlike other antibodies (e.g., anti-PD-1, anti-PD-L1, anti-CTLA-4), which have well-characterized dosing regimens for mouse models, clone 9E9 does not have such published standards.
• Strain-specific adjustments: While no major inter-strain differences are reported, researchers should empirically test and optimize dosing in their specific mouse model, as is standard practice for antibodies without established protocols.
• Consult literature and vendors: For the most reliable results, contact the antibody supplier and review any application notes or tech support documents. Additionally, a thorough literature search for recent papers using clone 9E9 in vivo may yield practical dosing examples.

Summary Table

Conclusion

Dosing regimens for clone 9E9 are not well established and do not appear to vary substantially across mouse strains based on current evidence. Researchers should determine optimal dosing empirically for their specific model and application, and consult both the antibody supplier and the scientific literature for any emerging data.

If you require precise dosing, consider reaching out to laboratories with published experience using clone 9E9 in your model of interest, or perform pilot dose-ranging experiments.