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

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

Product No.: C3346

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Clone
MJ18
Target
CD276
Formats AvailableView All
Product Type
Monoclonal Antibody
Alternate Names
CD276; CD276 antigen; B7h3; B7RP-2; AU016588; 6030411F23Rik
Isotype
Rat IgG1 κ
Applications
B
,
FC
,
in vivo

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Antibody Details

Product Details

Reactive Species
Mouse
Host Species
Rat
Recommended Isotype Controls
Recommended Dilution Buffer
Immunogen
Mouse B7-H3 IgG2a fusion protein
Product Concentration
≥ 5.0 mg/ml
Endotoxin Level
<1.0 EU/µg as determined by the LAL method
Purity
≥95% monomer by analytical SEC
>95% by SDS Page
Formulation
This monoclonal antibody is aseptically packaged and formulated in 0.01 M phosphate buffered saline (150 mM NaCl) PBS pH 7.2 - 7.4 with no carrier protein, potassium, calcium or preservatives added. Due to inherent biochemical properties of antibodies, certain products may be prone to precipitation over time. Precipitation may be removed by aseptic centrifugation and/or filtration.
Product Preparation
Functional grade preclinical antibodies are manufactured in an animal free facility using in vitro cell culture techniques and are purified by a multi-step process including the use of protein A or G to assure extremely low levels of endotoxins, leachable protein A or aggregates.
Storage and Handling
Functional grade preclinical antibodies may be stored sterile as received at 2-8°C for up to one month. For longer term storage, aseptically aliquot in working volumes without diluting and store at ≤ -70°C. Avoid Repeated Freeze Thaw Cycles.
Country of Origin
USA
Shipping
Next Day 2-8°C
Applications and Recommended Usage?
Quality Tested by Leinco
FC
Additional Applications Reported In Literature ?
B
FC
Each investigator should determine their own optimal working dilution for specific applications. See directions on lot specific datasheets, as information may periodically change.

Description

Description

Specificity
MJ18 activity is directed against mouse CD276 also known as B7-H3.
Background
CD276, also known as B7 homolog 3 protein (B7-H3), is a member of the B7 superfamily and acts as an immune checkpoint molecule and a costimulatory/coinhibitory immunoregulatory protein1. CD276 influences innate and adaptive immunity, regulates the aggressiveness of cancer cells, and is thought to play an important role in tumor development and cancer immunity. CD276 has been studied in many cancers, including breast, lung, ovarian, brain, gastric, and squamous cell carcinoma.

Human CD276 exists as either a soluble isoform or as a ~45–66 kDa type I transmembrane protein that is composed of an extracellular domain, a transmembrane domain, and a short intracellular domain1. In murine CD276, the extracellular domain is composed of a single pair of immunoglobulin variable and constant domains. Soluble CD276 is produced by cleavage from the cell surface or via alternative intron splicing and has been found in the secretomes of exosomes and other extracellular vesicles.

In normal human tissues, CD276 mRNA is widely and abundantly expressed but protein abundance is low1. miR-124 is thought to cause translational repression of CD276 by targeting its 3’-UTR, while other miRNAs are known to affect CD276 expression. In contrast, in tumor cells, CD276 mRNA and protein are abundant, and its presence is correlated with worsened prognosis, poor survival, recurrence rate, and enhanced invasive and migratory properties. CD276 blocking with monoclonal antibodies has been shown to reduce tumor growth and prolong survival in mouse models of various cancers. TREM-like transcript 2 (TLT-2) has been identified as a potential receptor2.

MJ18 was generated by immunizing Sprague Dawley rats with aa 1–242 of mouse CD276 extracellular domain linked to the Fc portion of mouse IgG2a3. After immunization, lymph node cells were fused with P3U1 myeloma cells, hybridomas were selected by flow cytometry, and MJ18 was purified from ascites.
Antigen Distribution
CD276 is weakly expressed on activated lymphocytes, macrophages, dendritic cells, nasal and airway epithelial cells, osteoblasts, and some tumor cell lines. A soluble form is secreted by monocytes, dendritic cells, and activated T cells.
NCBI Gene Bank ID
UniProt.org
Research Area
Immunology

Leinco Antibody Advisor

Powered by AI: AI is experimental and still learning how to provide the best assistance. It may occasionally generate incorrect or incomplete responses. Please do not rely solely on its recommendations when making purchasing decisions or designing experiments.

Clone MJ18 is commonly used in vivo in mice for applications aimed at blocking the function of CD276 (B7-H3), an immune checkpoint molecule thought to dampen T cell activity and promote tumor immune evasion. The intended uses have included:

  • Blocking B7-H3 signaling to study its role in immune suppression and tumor immunity.
  • Investigating tumor growth, tumor regression, and immune cell infiltration in syngeneic and xenograft mouse tumor models, especially to assess the impact of B7-H3 blockade on antitumor immune responses.
  • Exploring T cell function and activation in vivo by interfering with negative immune regulatory pathways mediated by B7-H3.

Critical context:
Recent, rigorous studies have questioned the specificity of clone MJ18 for B7-H3 in mice. While many published studies used MJ18 under the assumption it blocks murine B7-H3, newer evidence demonstrates MJ18 does not bind B7-H3 on murine tumor cell lines, but instead appears to interact with Fc receptors on the surface of murine immune cells—especially B cells and some T cells.
Thus:

  • Reported in vivo applications include tumor immunotherapy experiments and immune modulation studies, but the true molecular target of MJ18 in mice is likely not B7-H3, making earlier interpretations potentially invalid.
  • Validation with alternative, confirmed anti-B7-H3 clones (such as EPNCIR122) is recommended when studying murine B7-H3 biology.

Summary of reported in vivo uses:

  • Blocking (putative) B7-H3 signaling on immune/tumor cells.
  • Modulation of immune cell activity in cancer and immunology models.
  • Assessment of tumor growth/regression and immune infiltration after antibody administration in murine cancer models.
  • Flow cytometry or functional blockade experiments to probe immune checkpoint pathways.

Important limitation:
Given the lack of validated B7-H3 specificity for MJ18 in mouse in vivo systems, results from studies using MJ18 should be interpreted with caution, and researchers are encouraged to seek independent validation.

In the scientific literature, several antibodies and proteins are commonly used alongside MJ18. These include:

  • EPNCIR122 (EPN): This is another antibody specific for B7-H3, often used for comparative studies. It is noted for its specificity in targeting murine and human B7-H3, which contrasts with the non-specific binding observed for MJ18.

  • NoAb (no antibody control): This is used as a control in experiments to ensure that any observed effects are due to the antibody of interest (e.g., MJ18) rather than non-specific interactions.

  • Fc receptor proteins: MJ18 has been found to bind to Fc receptors on the surface of murine splenocytes, which complicates its use as a specific B7-H3 targeting agent.

These proteins and antibodies are utilized in various research contexts, including immunological studies and cancer research, to better understand immune responses and potential therapeutic targets.

The key findings from scientific literature regarding clone MJ18 are that MJ18, although widely cited as a blocking antibody for murine B7-H3 (CD276), actually does not bind B7-H3, but instead recognizes the Fc receptor FcγRIIB on murine splenocytes, predominantly on B cells.

Essential context and supporting details:

  • MJ18 does not recognize B7-H3: Multiple studies have demonstrated through flow cytometry and immunoprecipitation-mass spectrometry (IP-MS) that MJ18 does not bind to B7-H3 on mouse tumor cells or splenocytes, contradicting prior assumptions and product descriptions.
  • Recognizes FcγRIIB: MJ18 binds to the FcγRIIB receptor on murine B cells and T cells, rather than B7-H3, as validated by reduced fluorescence when Fc receptors are blocked.
  • Implications for antibody use: The literature highlights an erroneous use of MJ18 as a B7-H3-blocking antibody, advising that its targeting mechanism should be re-evaluated in studies where MJ18 was assumed to inhibit B7-H3. Validation using FcγRIIB knockout mice is suggested to conclusively confirm this target.
  • Prior product claims: Despite some commercial sources stating that MJ18 “reacts with mouse CD276 (B7-H3)” and has been used to inhibit CD4+ T-cell proliferation, recent peer-reviewed research has shown that these applications may be based on incorrect assumptions about epitope recognition.

Additional relevant information:

  • Historical use: Previous studies used MJ18 in tumor immunology models and cited its function as a B7-H3 inhibitor, with experimental protocols based on this understanding.
  • Validated B7-H3 antibody: The clone EPNCIR122 (distinct from MJ18) has been validated as a specific antibody for murine and human B7-H3 detection and blockade.
  • Recommendation: Researchers are advised to reassess the results and interpretations of experiments involving MJ18 as a surrogate for B7-H3 blockade, given its actual binding specificity.

In summary, MJ18 does not bind B7-H3 and instead targets FcγRIIB, underscoring a significant correction to past scientific practice and literature regarding its use.

Based on available evidence, dosing regimens of clone MJ18 show minimal variation across different mouse models, with researchers typically applying a standardized protocol regardless of the specific experimental context.

Standard Dosing Protocol

The most commonly reported dosing regimen for MJ18 is remarkably consistent: 300 μg per injection administered every other day via intraperitoneal injection. This protocol has been adopted as the standard approach across various experimental settings, including genetically modified mouse models and syngeneic tumor systems.

For example, in studies using Tgfbr1/Pten 2cKO mice (a genetically engineered model with mixed FVBN/CD1/129/C57 background), researchers administered 0.3 mg of MJ18 intraperitoneally every other day starting from day 14 after tamoxifen induction. Similarly, in orthotopic rhabdomyosarcoma models (M3-9-M), investigators used the identical 300 μg per injection every other day regimen.

Lack of Model-Specific Variations

No significant published evidence suggests major variations of the MJ18 dosing regimen based on different mouse strains or tumor models. This uniformity persists despite differences in:

  • Genetic backgrounds (pure strains versus mixed backgrounds)
  • Tumor types and growth kinetics
  • Immunogenicity of the tumor models
  • Treatment initiation timing

The standardized approach contrasts with the model-specific optimization often seen with other checkpoint blockade antibodies, where doses may range significantly (for example, anti-PD-1 antibodies typically use 100-500 μg per mouse depending on the clone and application).

Important Caveat

Recent research has revealed that MJ18 does not actually recognize B7-H3 on the surface of mouse cells, despite being widely used as an anti-B7-H3 blocking antibody. Instead, MJ18 appears to bind Fc receptors on immune cells, particularly B cells. This finding raises questions about the mechanistic basis for any observed effects and suggests that the uniform dosing regimen may reflect convention rather than optimized targeting of a specific antigen.

References & Citations

1. Zhou WT, Jin WL. Front Immunol. 12:701006. 2021.
2. Hashiguchi M, Kobori H, Ritprajak P, et al. Proc Natl Acad Sci USA. 105: 10495–10500. 2008.
3. Nagashima O, Harada N, Usui Y, et al. J Immunol. 181(6):4062-71. 2008.
4. Yamato I, Sho M, Nomi T, et al. Br J Cancer. 101(10):1709-1716. 2009.
5. Kamachi F, Isshiki T, Harada N, et al. Biochem Biophys Res Commun. 463(4):739-45. 2015.
B
Flow Cytometry
in vivo Protocol

Certificate of Analysis

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Formats Available

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Disclaimer AlertProducts are for research use only. Not for use in diagnostic or therapeutic procedures.