Anti-Mouse CD276 (Clone MJ18) – Purified in vivo PLATINUM™ Functional Grade

Common In Vivo Applications of Clone MJ18 in Mice

Clone MJ18 is a rat anti-mouse monoclonal antibody raised against the immune checkpoint molecule CD276 (B7-H3). It has been widely used in mouse research, especially in the fields of immunology and immuno-oncology.

Reported Applications

• Blocking B7-H3 (CD276) signaling in vivo: MJ18 is primarily used to block the function of B7-H3 on immune cells and tumor cells in mice, with the goal of modulating immune responses and enhancing antitumor immunity.
• Studying immune response modulation: By inhibiting B7-H3 signaling, MJ18 is employed to investigate how this checkpoint regulates T cell activation, proliferation, and immune tolerance. The rationale is that B7-H3 acts as a negative regulator of T cell responses, and its blockade may promote immune activation.
• Tumor microenvironment research: MJ18 has been used in syngeneic and orthotopic mouse tumor models to assess the impact of B7-H3 blockade on tumor growth, immune cell infiltration, and tumor regression. Some studies have reported tumor regression and increased T cell infiltration with MJ18 treatment, although these findings are controversial (see below).
• In vivo flow cytometry and immune phenotyping: MJ18 is also used in flow cytometry to identify B7-H3-expressing subsets in mouse tissues, although its specificity has recently come under scrutiny.

Controversies and Challenges

Recent data suggest that MJ18 may not specifically bind mouse B7-H3 on tumor cells but instead targets unidentified ligands on murine splenocytes, particularly Fc receptors (e.g., FcγRIIB). Importantly, in a rhabdomyosarcoma mouse model, MJ18 failed to induce tumor regression as would be expected if it targeted B7-H3, whereas B7-H3 knockout models showed delayed tumor growth. This raises significant concerns about the validity of previous studies using MJ18 for B7-H3 blockade in mice.

Furthermore, technical validations (flow cytometry and immunoprecipitation-mass spectrometry) indicate that MJ18 binds splenocytes independently of B7-H3, with FcγRIIB identified as a probable target—an inhibitory Fc receptor expressed on B cells and some T cells. These findings suggest that observed immunological effects attributed to MJ18 in vivo may actually result from its interaction with Fc receptors, not B7-H3.

Summary Table: Reported vs. Critically Assessed Uses

Conclusion

While clone MJ18 is widely cited and sold for in vivo mouse studies targeting B7-H3, recent rigorous validation studies strongly question its specificity and mechanistic basis for B7-H3 blockade. It remains a common tool in preclinical research for modulating immune responses and tumor immunity, but its results should be interpreted with caution and ideally confirmed with alternative, validated antibodies (such as EPNCIR122). Researchers are encouraged to validate MJ18’s binding and functional effects in their specific experimental systems and consider the possibility of off-target, Fc receptor-mediated interactions.

Other commonly used antibodies or proteins paired with MJ18 in the literature include EPNCIR122 (EPN), NoAb (no antibody control), and various Fc receptor proteins for comparative studies and controls.

Significant details from the literature:

• EPNCIR122 (EPN): This antibody is validated as specific for both murine and human B7-H3 and is often used as a critical comparator when investigating MJ18 binding specificity.
• NoAb (No antibody control): Used to identify baseline or background binding in experiments, aiding in the assessment of MJ18's specific vs. nonspecific interactions.
• Fc receptor proteins: Fcγ receptors (e.g., FcγRI, FcγRIIB, FcγRIII, FcγRIV, FcRn) have been found as MJ18 binders in murine splenocytes, suggesting cross-reactivity or binding unrelated to B7-H3/CD276.
• LCP1 (Lymphocyte Cytosolic Protein 1): Identified in MJ18 pulldown samples, though subsequent validation indicates MJ18 binding occurs even in LCP1-deficient mice, suggesting it is not the universal target.

MJ18 is often used in conjunction with these reagents in assays such as immunoprecipitation, flow cytometry, and immunophenotyping, especially when studying immune cell populations or validating antibody specificity.

In summary, EPNCIR122 (EPN), NoAb controls, and Fc receptor proteins are the most commonly reported alongside MJ18 in recent studies, mainly for specificity validation and control purposes.

Clone MJ18, initially reported as an anti-mouse CD276 (B7-H3) antibody, has been a subject of conflicting findings in scientific literature. Here are the key points:

1. Misidentification as B7-H3 Antibody: Contrary to earlier claims, MJ18 does not bind to B7-H3. Studies have shown that MJ18 does not recognize murine B7-H3 but instead binds to the Fc receptor FcγRIIB on murine splenocytes.

2. Immune Modulation: Despite not targeting B7-H3, MJ18 has been used in studies aimed at modulating immune responses. Early reports suggested that MJ18 could enhance T cell infiltration and cause tumor regression, although these effects were not due to blocking B7-H3.

3. Fc Receptor Interaction: The primary interaction of MJ18 is with Fc receptors, particularly FcγRIIB, which is expressed on B cells and some T cells. This interaction may influence immune cell function.

4. Potential Applications: While MJ18's effects on immune modulation need further clarification, it remains a tool for studying immune responses in preclinical settings.

In summary, the scientific community has revised its understanding of MJ18's target, and ongoing research aims to explore its actual biological effects.

The dosing regimens for the MJ18 antibody, targeted against mouse CD276 (B7-H3), generally follow a consistent pattern across different mouse models. The most commonly reported dosing regimen for MJ18 is 300 μg per injection every other day. This protocol has been utilized in various studies, including those involving tumor models, where it aims to block the B7-H3 interaction, although recent findings suggest that MJ18 may not actually bind to B7-H3 as previously thought.

In specific mouse models, such as the Tgfbr1/Pten 2cKO mice, the same dosing of 0.3 mg (300 μg) of MJ18 every other day has been applied. While there is no significant variation in dosing based on different mouse strains or tumor models, the specific dosing may depend on the experimental design and the goals of the study. For example, factors like the desired level of immune modulation or the specific tumor type might influence the choice of dosing regimen, but variation from the standard protocol is not commonly reported.

Key Points:

• Standard Dose: 300 μg per injection every other day.
• Consistency Across Models: This regimen is generally consistent across different mouse models.
• Experimental Variability: Specific experimental conditions may lead to variations in dosing, but such variations are not widely documented.