Recombinant Mouse CCL22

Recombinant Mouse CCL22 is used in research to study immune cell trafficking, tumor immunology, metabolic regulation, and inflammatory diseases due to its role as a chemokine that attracts specific immune cell subsets, particularly those expressing the CCR4 receptor.

Key scientific applications and rationale include:

• Immune Cell Chemotaxis and Trafficking: CCL22 is a potent chemoattractant for CCR4-expressing cells, such as Th2 cells and regulatory T cells (Tregs), making recombinant CCL22 valuable for in vitro chemotaxis assays, migration studies, and dissecting mechanisms of immune cell recruitment to sites of inflammation or tumors.

• Tumor Microenvironment and Cancer Immunology: CCL22 is implicated in the recruitment of Tregs to the tumor microenvironment, which can suppress anti-tumor immunity. Recombinant CCL22 can be used to model or manipulate this process in vitro or in vivo, aiding studies on tumor immune evasion and the development of immunotherapies targeting the CCL22-CCR4 axis.

• Metabolic and Thermogenic Research: Recent studies show that exogenous recombinant CCL22 can promote adipose tissue beiging (conversion of white to beige fat), increase energy expenditure, and reduce fat mass in mice, highlighting its utility in metabolic and obesity research. Supplementation with recombinant CCL22 restored thermogenic gene expression and promoted adaptive thermogenesis in mouse models.

• Inflammatory and Allergic Disease Models: CCL22 is highly expressed in lesions of T cell-mediated inflammatory diseases (e.g., atopic dermatitis, asthma, psoriasis). Recombinant CCL22 can be used to study the mechanisms of immune cell infiltration and cytokine regulation in these contexts.

• Functional Assays: Recombinant CCL22 is used to stimulate or modulate immune cells in vitro, such as inducing calcium mobilization or chemotaxis in T cells, and to study downstream signaling pathways.

• Mechanistic Studies: By adding recombinant CCL22 to cell cultures or animal models, researchers can dissect the specific roles of the CCL22-CCR4 axis in immune regulation, tissue homeostasis, and disease pathogenesis.

Summary of best practices:

• Use recombinant CCL22 in controlled in vitro assays to study chemotaxis, signaling, or gene expression changes in CCR4+ cells.
• Employ in vivo administration to model immune cell recruitment, metabolic effects, or to test therapeutic interventions targeting the CCL22-CCR4 pathway.
• Validate specificity using CCR4-deficient cells or animals as controls.

In summary, recombinant mouse CCL22 is a versatile tool for dissecting immune cell dynamics, tumor immunology, metabolic regulation, and inflammatory disease mechanisms in both in vitro and in vivo systems.

Yes, recombinant mouse CCL22 can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated for your specific assay system.

Recombinant mouse CCL22 is commonly used as a standard in commercial ELISA kits designed to quantify mouse CCL22 (also known as MDC) in biological samples. These kits typically include a recombinant CCL22 standard, and the assay is validated to ensure that the standard curve generated with recombinant protein is parallel to the response obtained with native CCL22 in samples.

Key considerations:

• Parallelism: Commercial kits demonstrate that standard curves generated with recombinant CCL22 are parallel to those obtained with natural CCL22, indicating that the recombinant protein is suitable for accurate quantification.
• Matrix Effects: When using recombinant CCL22 as a standard, ensure that the diluent used for the standard curve matches the sample matrix as closely as possible to minimize matrix effects. Some kits recommend evaluating diluents for complex matrices like serum or plasma.
• Validation: If you are developing your own ELISA or using a kit for a novel application, validate that your recombinant CCL22 standard produces a linear and parallel standard curve compared to native CCL22 in your sample type.
• Storage and Handling: Follow best practices for reconstitution and storage of recombinant standards, typically at -20 °C or -80 °C after reconstitution, to maintain protein integrity.

Summary of best practices:

• Use recombinant mouse CCL22 as a standard if your ELISA is designed for mouse CCL22 and the antibodies used recognize both recombinant and native forms.
• Validate parallelism and recovery in your specific assay and sample matrix.
• Prepare the standard curve using the same buffer or matrix as your samples when possible to ensure accurate quantification.

If you are using a commercial ELISA kit, follow the manufacturer’s protocol for standard preparation and curve fitting. If developing your own assay, ensure thorough validation with both recombinant and native CCL22.

Recombinant Mouse CCL22 has been validated for several key applications in published research, primarily in studies of immunology, cancer, metabolism, and inflammation.

Validated Applications:

• Bioactivity Assays: Recombinant Mouse CCL22 is routinely validated for bioactivity, including its ability to induce chemotaxis and calcium mobilization in activated mouse and human T cells. This is a standard functional assay to confirm the protein's activity.

• ELISA Standards and Controls: Recombinant CCL22 is used as a standard or control in ELISA assays to quantify CCL22 levels in biological samples such as cell culture supernatants, serum, plasma, and tissue homogenates. This enables accurate measurement of endogenous CCL22 in various experimental contexts.

• In Vivo Functional Studies: Recombinant CCL22 has been administered to mice to study its physiological effects, such as promoting beiging of inguinal white adipose tissue (iWAT) in response to cold exposure, and recruiting eosinophils to adipose tissue. These studies use recombinant protein injections to directly assess CCL22's role in metabolic regulation.

• Immunomodulation and Cancer Research: CCL22 is central to studies investigating its role in recruiting regulatory T cells (Tregs) to the tumor microenvironment, modulating immune cell infiltration, and influencing anti-tumor immunity. Recombinant CCL22 is used to probe these mechanisms in vitro and in vivo.

• Autoimmunity and Inflammation Models: Recombinant CCL22 has been used in models of autoimmune diabetes to study Treg recruitment to pancreatic islets, and in pulmonary fibrosis to investigate macrophage activation and immune cell trafficking.

Supporting Details:

• Chemotaxis Assays: The protein is validated for its ability to chemoattract Th2 cytokine-producing cells and eosinophils, confirming its functional relevance in immune cell migration studies.

• Cellular and Molecular Mechanism Studies: Recombinant CCL22 is used to dissect signaling pathways, such as CCR4-mediated responses, and to study its effects on gene expression (e.g., UCP1 induction in adipocytes).

• Disease Models: Published research has employed recombinant CCL22 in models of cancer, autoimmune disease, pulmonary fibrosis, and metabolic regulation to elucidate its biological roles and therapeutic potential.

Summary Table of Validated Applications

Conclusion:
Recombinant Mouse CCL22 is a well-validated tool for studying chemokine biology, immune cell trafficking, metabolic regulation, and disease mechanisms in both in vitro and in vivo settings. Its applications span bioactivity assays, ELISA, functional studies, and disease modeling, making it integral to immunological and biomedical research.

To reconstitute and prepare Recombinant Mouse CCL22 protein for cell culture experiments, add sterile water or sterile PBS to the lyophilized protein to achieve a stock concentration between 0.1–1.0 mg/mL. Avoid vigorous mixing (such as vortexing) and do not exceed 1 mg/mL during reconstitution.

Step-by-step protocol:

• Reconstitution:

  • Add sterile distilled water or sterile PBS to the vial to reach the desired stock concentration (commonly 0.1–1.0 mg/mL).
  • Gently mix by pipetting up and down or by slow inversion. Do not vortex.
  • If the formulation contains excipients (e.g., trehalose, mannitol), these will dissolve upon reconstitution.

• Dilution for cell culture:

  • Prepare working solutions by diluting the stock in cell culture medium or buffer containing a carrier protein (e.g., 0.1% BSA or serum albumin) to minimize adsorption and loss of activity.
  • Typical working concentrations for chemotaxis or functional assays range from 0.5–20 ng/mL, depending on cell type and experimental design.

• Storage:

  • Store reconstituted protein at 4°C for short-term use (2–7 days).
  • For long-term storage, aliquot and freeze at −20°C or −80°C. Avoid repeated freeze-thaw cycles.
  • If using carrier-free protein, consider adding a carrier protein for long-term stability.

Additional notes:

• Confirm the protein’s bioactivity and endotoxin level if required for sensitive cell types.
• Always use sterile technique to prevent contamination.
• If the protein is provided in a formulation with excipients (e.g., trehalose, mannitol), these do not interfere with most cell culture applications.

Summary Table: Preparation and Storage

This protocol ensures optimal solubility, stability, and activity of recombinant mouse CCL22 for cell culture experiments.