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

Clone 9E9 is used in in vivo mouse studies primarily to block mouse Fc?RIV (CD16.2), and also Fc?RIII, in order to study the functional roles of these receptors in immune responses, such as platelet clearance and immune complex handling.

In experimental designs, 9E9 is administered to mice to selectively inhibit Fc?RIV (CD16.2), an Fc gamma receptor expressed on myeloid cells including monocytes, macrophages, dendritic cells, and neutrophils—but not lymphoid cells (B, T, NK cells). This blocking allows researchers to:

• Assess the necessity of Fc?RIV for IgG2a- and IgG2b-mediated platelet clearance, confirming that Fc?RIV is required for these processes in vivo.
• Investigate the roles of Fc?RIV and Fc?RIII in immune cell activity, since 9E9 can block both Fc?RIV and, in certain contexts, Fc?RIII (when 9E9 is first bound to Fc?RIV on the same cell).
• Reduce B cell depletion mediated through these receptors, giving insight into the mechanism of antibody-mediated immune cell clearance.
• Interfere with immune complex binding to Fc?RIV, helping to clarify mechanisms of immune complex processing.

Experimental applications include:

• Blocking/neutralizing receptor function during antibody therapy, autoimmune disease models, or immunology studies.
• Tracking changes in immune cell behavior when Fc?RIV is blocked.
• Distinguishing the contributions of individual Fc gamma receptor subtypes in vivo.

Key experimental features:

• Clone 9E9 is a monoclonal antibody from Armenian hamster IgG, used at concentrations determined by specific protocols (commonly 1–5 mg/ml).
• The antibody does not activate the receptor but blocks ligand binding and downstream effects.
• It is supplied in a sterile, preservative-free PBS solution, certified low-endotoxin for sensitive in vivo studies.

In summary, clone 9E9 is leveraged as a functional-blocking reagent for Fc?RIV (and secondary Fc?RIII blockade) during in vivo studies to dissect the biology of IgG-mediated immune responses and Fc receptor functions in mice.

The correct storage temperature for sterile packaged clone 9E9 is 4°C (refrigerator temperature), and it should not be frozen.

This storage condition applies to the stock concentration and is maintained to preserve sterility and antibody integrity. Supporting information from other protocols involving sterile antibody solutions indicates that refrigeration at 4°C is standard practice as long as sterility is not compromised. Freezing of the product is explicitly discouraged, as this may damage the antibody and compromise its functionality.

The antibody 9E9 is most commonly used to target mouse Fc?RIV (CD16.2), and in studies using 9E9, other frequently used antibodies or proteins include those targeting related Fc gamma receptors and immune markers:

• Anti-mouse Fc?RII (CD32)
• Anti-mouse Fc?RIII (CD16)
• Isotype controls (often Armenian Hamster IgG, due to the host species of 9E9)
• Detection antibodies for cell markers, such as CD11b (myeloid cells), Ly6G (neutrophils), F4/80 (macrophages), and CD45 (leukocytes) in flow cytometry and immunophenotyping
• Antibodies against immunoglobulin subclasses (e.g., anti-mouse IgG2a, IgG2b), especially for studies on platelet clearance and antibody-dependent cell depletion
• Other anti-Fc?R antibodies (e.g., anti-Fc?RI/CD64, anti-Fc?RIIb/CD32B), often used in multiplex panels to distinguish roles of different Fc? receptors in immune function

These antibodies and proteins are employed for functional studies, blocking experiments, flow cytometry, and Western blotting to dissect Fc?R signaling, immune cell depletion, and antibody-dependent processes. In some research, a sandwich assay using secondary anti-Armenian hamster IgG antibodies is used for quantitative detection since 9E9 is an Armenian hamster monoclonal and species-specific detection kits may not be compatible.

In summary, 9E9 is frequently paired with other Fc?R-targeting antibodies, lineage markers, and immunoglobulin detection tools to interrogate the functional interplay of immune receptors and cells, especially in mouse immunology and antibody clearance studies.

The key findings from scientific literature citations on clone 9E9 show that it is an anti-Fc?RIV (Fc gamma receptor IV) monoclonal antibody used primarily to block or deplete Fc?RIV function in mouse immune cells.

Key points supported by the literature:

• Specific Inhibition of Fc?RIV: Treatment with the anti-Fc?RIV clone 9E9 antibody leads to a significant reduction of Fc?RIV expression on the surface of all splenic antigen-presenting cell subsets analyzed. This demonstrates its effectiveness as a blocking antibody in experimental models where Fc?RIV functions are dissected.

• Broader Impacts on Fc? Receptors: Administration of clone 9E9 unexpectedly also reduces surface levels of Fc?RIII and Fc?RII (but not Fc?RI) on different immune cell subsets. The literature notes that these off-target effects should be considered when interpreting experimental outcomes using 9E9.

• Experimental Utility in Disease Models: Studies have used clone 9E9 to show that blocking Fc?RIV can ameliorate autoimmune and alloimmune pathologies mediated by IgG2a/c antibodies (such as in models of autoimmune hemolytic anemia, rheumatoid arthritis, and nephritis). Therefore, Fc?RIV—and by extension clone 9E9—is considered a potential therapeutic target in these diseases.

• Mechanistic Clarifications: Extensive investigation found that despite the reduction in Fc?RIII and Fc?RII expression upon 9E9 treatment, their presence or absence does not confound the primary effect, as knockout mice for Fc?RIII or the FcR? chain did not recapitulate the phenotype observed with 9E9, confirming the specificity of 9E9-mediated Fc?RIV blockade in the studied immune responses.

• Application in Immunological Research: Clone 9E9 is thus a widely used tool for functional studies of Fc?RIV in vivo, commonly utilized in flow cytometry, depletion, or blocking experiments in mice.

In summary, clone 9E9 is recognized as a well-validated, highly specific monoclonal antibody tool to interrogate the biology of Fc?RIV in mouse models, with broad application in autoimmunity, alloimmunity, and basic immunology research.