CD44 Mouse Antibody

Rat Anti-Mouse CD44
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Description

Key CD44 Mouse Antibody Clones and Characteristics

The following table summarizes prominent CD44 mouse antibody clones, their properties, and applications:

CloneIsotypeSpecificityApplicationsCross-Reactivity
IM7IgG2bAll CD44 isoformsFlow cytometry, CDC assays, immunohistochemistryMouse, human, dog, cat, horse
5035-41.1DIgG1CD44 (multiple strains)Flow cytometry, immunoprecipitationC57BL/6, AKR, C3H, NZB mice
C44Mab-46IgG1CD44 standard/variant isoformsFlow cytometry, Western blot, xenograft therapyHuman, mouse
8E2IgG1Total CD44Western blot, immunofluorescenceHuman, rat
156-3C11IgG2aCD44 (human-specific)Western blot, immunohistochemistryHuman

Antitumor Activity in Xenograft Models

  • C44Mab-46-mG2a and 5-mG2a demonstrated potent antitumor effects in esophageal cancer models:

    • Induced 67–88% cytotoxicity in CHO/CD44s cells and 27–31% cytotoxicity in KYSE770 cells via complement-dependent cytotoxicity (CDC) .

    • Suppressed tumor growth by >50% compared to controls in xenograft models .

Diagnostic and Functional Assays

  • IM7:

    • Recognizes hematopoietic and non-hematopoietic cells, including memory T cells .

    • Used in flow cytometry at ≤0.125 µg/test for murine bone marrow and splenocytes .

  • C44Mab-46:

    • Detects CD44s in Western blot (apparent KD: 1.1 × 10⁻⁸ M for CHO/CD44s) .

    • Stained formalin-fixed esophageal squamous cell carcinoma tissues effectively .

Complement-Dependent Cytotoxicity (CDC)

  • 5-mG2a and C44Mab-46-mG2a activated effector cells and mediated CDC against CD44-positive tumors :

    AntibodyCHO/CD44s CytotoxicityKYSE770 Cytotoxicity
    5-mG2a67%27%
    C44Mab-46-mG2a88%31%

Anti-Inflammatory Effects

  • Anti-CD44 antibodies (e.g., IM7) block FcγR-mediated phagocytosis in macrophages, reducing platelet clearance in immune thrombocytopenia .

  • Deglycosylated or F(ab′)₂ variants lose therapeutic efficacy, confirming Fc-dependent mechanisms .

Challenges and Future Directions

  • Dose Optimization: Preclinical studies highlight the need for precise dosing to balance tumor targeting and off-tissue effects .

  • Isoform-Specific Targeting: Variant isoforms (e.g., CD44v3–10) may require tailored antibodies for cancer stem cell eradication .

Product Specs

Introduction
CD44 is a cell surface glycoprotein involved in cell-cell interactions, adhesion, and migration. It acts as a receptor for hyaluronic acid and interacts with other ligands like osteopontin, collagens, and matrix metalloproteinases. CD44 participates in various cellular functions, including lymphocyte activation, recirculation, homing, hematopoiesis, and tumor metastasis. In multiple sclerosis, CD44 and CD49d are considered potential activity markers, with CD44 as a possible therapeutic target. Increased CD44 antigen is linked to relapses in non-small cell lung cancers.
Formulation
Supplied at a concentration of 1 mg/ml in PBS buffer after reconstitution.
Storage Procedures
The lyophilized antibody should be stored at 4°C. After reconstitution, use within one month. For long-term storage, aliquot the reconstituted antibody and store at -20°C.
Solubility
Reconstitute the lyophilized antibody with sterile H2O. Mix gently by inverting the vial several times. Rinse the sides of the vial and allow 30-60 seconds for complete reconstitution before use.
Applications
This antibody is suitable for staining and immunohistochemistry applications. For staining, a recommended concentration is 10 µl per 1,000,000 cells. The optimal titer for immunohistochemistry should be determined by the investigator.
Available Conjugates
This antibody is available in various conjugated forms, including biotin and FITC conjugates.
Synonyms
MDU2, MDU3, MIC4, CDW44, CSPG8, HCELL, HUTCH-I, Phagocytic glycoprotein I, PGP-1, Extracellular matrix receptor-III, ECMR-III, Hermes antigen, Hyaluronate receptor, Heparan sulfate proteoglycan, Epican) CDw44.
Purification Method
Protein A.
Type
Rat Anti Mouse Monoclonal.
Clone
mCD44.
Immunogen
CD44 enriched mouse LN T cells.
Ig Subclass
Rat IgG.

Q&A

What critical experimental parameters determine CD44 antibody performance in murine models?

CD44 antibody efficacy depends on three factors: (1) clone-epitope specificity, (2) administration protocol, and (3) endpoint validation. The IM7 clone (binds conserved hyaluronan-binding domain) demonstrates superior in vivo neutralization compared to KM114 in metabolic studies, requiring daily 50-100 μg intraperitoneal doses for sustained CD44 shedding . Validation should include:

Validation MetricIM7 Clone Performance KM114 Clone Performance
In vivo neutralizationReduces CD44+ leukocytes by 68%Limited neutralization data
Inflammatory marker suppressionTNF-α ↓72%, IL-6 ↓65%Not characterized
Metabolic impactFasting glucose ↓40%No reported metabolic effects

Endpoint analysis must quantify both membrane-bound CD44 (flow cytometry) and shed soluble CD44 (ELISA), as certain clones preferentially detect either form .

How does CD44 isoform complexity impact antibody selection?

CD44 exists in ≥19 isoforms through alternative splicing . While IM7 recognizes all isoforms due to its epitope location in the conserved hyaluronan-binding domain, clones like KM201 show isoform-specific binding patterns. Researchers must:

  • Perform isoform-specific RT-PCR on target tissues

  • Compare antibody binding affinity across isoforms using surface plasmon resonance

  • Validate detection specificity through CRISPR-mediated isoform knockout controls

In adipose tissue inflammation models, IM7’s pan-isoform neutralization proved essential for reducing macrophage infiltration (CD68+ cells ↓58%) compared to isoform-specific antibodies .

What mechanisms explain contradictory findings in CD44 antibody studies?

Contradictions arise primarily from four variables:

VariableImpact ExampleResolution Strategy
Mouse strainC57BL/6 vs. BALB/c show 3.2-fold CD44 expression differences Pre-screen CD44 expression via flow cytometry
Antibody timingEarly intervention (week 8) reduces glucose by 40% vs. 12% in late intervention (week 20) Standardize treatment windows
Disease modelHFD-induced diabetes vs. spontaneous T2D models show differential CD44 dynamicsCross-validate in ≥2 model systems
Endpoint selectionMembrane CD44 vs. soluble CD44 measurements yield opposing correlations with IRMulti-parametric analysis

A meta-analysis of 27 studies reveals that 68% of contradictions resolve when controlling for these variables .

How can researchers optimize CD44 antibody dosing for dual immune/metabolic endpoints?

Therapeutic dosing requires balancing immune modulation and metabolic effects:

ParameterImmune-OptimizedMetabolic-OptimizedDual-Action Protocol
Dose100 μg loading + 50 μg maintenance 25 μg continuous100 μg day 1 → 75 μg days 2-7 → 50 μg thereafter
FrequencyQDQODQD week 1 → Q3D maintenance
Outcome79% macrophage reduction35% glucose lowering62% macrophage reduction + 48% glucose lowering

This phased approach prevents CD44 receptor saturation while maintaining therapeutic levels, as validated in HFD models showing sustained HbA1c reductions (8.2% → 6.1%) over 12 weeks .

What orthogonal techniques resolve CD44 antibody specificity challenges?

Three confirmation methods are essential:

  • CRISPR-Cas9 CD44 knockout controls: Compare antibody signal in WT vs. KO tissues

  • Competitive binding assays: Pre-incubate with recombinant CD44 extracellular domain (10 μg/mL blocks >90% IM7 binding)

  • Multispectral imaging: Colocalize antibody signal with CD44-GFP reporters

In diabetes models, these methods revealed that 22% of commercial CD44 antibodies cross-react with CD74, necessitating rigorous validation .

How should researchers design controls for in vivo CD44 antibody studies?

A robust control panel includes:

Control TypeCompositionPurpose
IsotypeRat IgG2b κ Baseline immune activation
Ligand blockadeHyaluronan oligosaccharides (10 mg/kg) Confirm CD44-specific effects
GeneticCD44-/- miceAbsolute specificity verification
PharmacologicSoluble CD44-Fc fusion protein (1 mg/kg) Competitive inhibition control

In adipose tissue studies, this control matrix differentiated antibody-specific effects (68% inflammation reduction) from nonspecific immune activation (12% reduction in isotype controls) .

What biomarkers validate functional CD44 antibody activity in vivo?

Five essential biomarkers:

  • Soluble CD44: ↑2.4-fold in serum post-antibody administration

  • Adipose tissue CLS count: ↓ from 12.3 ± 1.2 to 5.1 ± 0.8 CLS/mm²

  • Hepatic CD44+ macrophages: Flow cytometry gating (CD45+CD11b+F4/80+CD44+)

  • Insulin signaling: p-AKT/AKT ratio in muscle and liver

  • Cytokine profile: Multiplex analysis of IL-1β, TNF-α, MCP-1

Longitudinal monitoring should occur at days 0, 7, 14, and 28 to capture dynamic responses .

Concluding Recommendations

  • Prioritize IM7 clone for metabolic studies requiring pan-isoform neutralization

  • Implement phased dosing regimens to balance efficacy and toxicity

  • Employ multi-parametric validation across ≥3 model systems

  • Standardize control matrices to isolate CD44-specific effects

Product Science Overview

Production and Characteristics

The Rat Anti-Mouse CD44 antibody is typically produced by immunizing rats with mouse CD44 antigen. The resulting hybridoma cells are then screened for the production of antibodies that specifically bind to CD44. These antibodies are purified from the culture supernatant or ascites fluid using affinity chromatography .

Some key characteristics of Rat Anti-Mouse CD44 antibodies include:

  • Isotype: Rat IgG2b, κ or Rat IgG2a, κ
  • Immunogen: Dexamethasone-induced, SJL mouse spontaneous myeloid leukemia M1 cells
  • Applications: Flow cytometry, immunohistochemistry (formalin-fixed paraffin and acetone-fixed frozen sections), and other immunoassays
Applications in Research

Rat Anti-Mouse CD44 antibodies are valuable tools in various research applications, including:

  1. Flow Cytometry: These antibodies are used to stain cells for the detection and quantification of CD44 expression on the cell surface. This is particularly useful for studying the activation and differentiation of immune cells .

  2. Immunohistochemistry: Rat Anti-Mouse CD44 antibodies are used to stain tissue sections to visualize the distribution and localization of CD44-expressing cells. This can provide insights into the role of CD44 in tissue architecture and pathology .

  3. Functional Studies: These antibodies can be used to block or stimulate CD44 function in various experimental settings. For example, they can be used to inhibit CD44-mediated cell adhesion or to induce signaling pathways downstream of CD44 .

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