Recombinant Mouse IL-31

Recombinant Mouse IL-31 is used in research applications to study its role as a cytokine involved in allergic reactions, inflammatory skin diseases, pruritus (itch), and immune regulation. It is particularly valuable for modeling disease mechanisms, testing therapeutic interventions, and dissecting cellular signaling pathways in mouse models.

Key scientific applications and rationale include:

• Modeling Pruritus and Dermatitis: Recombinant mouse IL-31 induces scratching behavior and skin inflammation in mice, making it a robust tool for studying the mechanisms of itch and dermatitis, especially in the context of atopic dermatitis and related skin disorders. This allows for the evaluation of anti-pruritic drugs and the investigation of the itch–scratch cycle.

• Immune Regulation Studies: IL-31 directly engages its receptor (IL-31RA/OSMRβ) on sensory neurons and immune cells, influencing both neurogenic inflammation and type 2 immune responses. It can restrain cutaneous type 2 inflammation via neuropeptide pathways, providing insight into the balance between inflammation and immune suppression in chronic skin diseases.

• Cellular Signaling and Proliferation: IL-31 affects cell proliferation in a cell type-dependent manner. For example, it stimulates proliferation of myeloid progenitor cells but suppresses lung epithelial cell proliferation, making it useful for studying cytokine signaling, cell cycle regulation, and hematopoietic homeostasis.

• Respiratory and Smooth Muscle Research: IL-31 and its receptor are implicated in airway inflammation and smooth muscle contraction, relevant for asthma and allergic airway disease models. Recombinant IL-31 can be used to investigate airway hyperresponsiveness and contractility in precision-cut lung slices and smooth muscle cell cultures.

• Therapeutic Target Validation: By administering recombinant IL-31 in vivo or in vitro, researchers can validate the efficacy of neutralizing antibodies or receptor antagonists, and explore novel therapeutic strategies for inflammatory and allergic diseases.

• Standardization and Reproducibility: Recombinant proteins provide a consistent, defined reagent for mechanistic studies, ensuring reproducibility across experiments and facilitating quantitative dose-response analyses.

In summary, recombinant mouse IL-31 is a critical reagent for dissecting the molecular and cellular mechanisms underlying pruritus, skin inflammation, immune modulation, and airway physiology in mouse models, and for preclinical testing of therapeutic interventions targeting IL-31 pathways.

Yes, recombinant mouse IL-31 can be used as a standard for quantification and calibration in ELISA assays. This is one of the primary applications of recombinant IL-31 proteins in research.

Characteristics of Recombinant Mouse IL-31 for ELISA Standards

Protein Specifications

Recombinant mouse IL-31 is typically expressed in Escherichia coli as a full-length protein spanning amino acids 24-163, with a molecular weight of approximately 15.7 kDa and containing 141 amino acids. The protein is produced as a non-glycosylated form with high purity (>95%) and low endotoxin levels (<0.1 EU/µg), making it suitable for quantitative applications.

Formulation and Storage

The recombinant protein is commonly supplied as a lyophilized powder at a concentration of 1 mg/mL in buffered solution (typically 10 mM sodium phosphate buffer, pH 7.5). For long-term storage and to maintain stability during reconstitution, it is recommended to add a carrier protein such as 0.1% human serum albumin (HSA) or bovine serum albumin (BSA). Lyophilized material remains stable at -20°C, while reconstituted material should be aliquoted and frozen at the same temperature.

Application in ELISA Workflows

Recombinant IL-31 serves as a calibration standard in sandwich ELISA assays designed to measure IL-31 in various sample types including serum, plasma, cell culture supernatants, and tissue homogenates. The sandwich ELISA principle relies on a matched antibody pair that captures the target between a pre-coated capture antibody and a detector antibody, with signal intensity directly proportional to target concentration. Using recombinant IL-31 as your standard allows you to generate accurate standard curves for quantifying both natural and recombinant IL-31 in your samples.

Quality Considerations

When using recombinant IL-31 as a standard, ensure that protein content and purity are determined through validated methods such as UV spectroscopy at 280 nm, reverse-phase high-performance liquid chromatography (RP-HPLC) calibrated against known standards, or quantitation via reducing and non-reducing SDS-PAGE gels. This verification ensures the accuracy of your standard curve and subsequent sample quantification.

Recombinant Mouse IL-31 has been validated for several applications in published research, primarily in the context of functional studies related to immunology, inflammation, and pruritus (itch).

Key validated applications include:

• Functional Assays: Recombinant mouse IL-31 is widely used to study its biological activity, such as inducing cytokine production (e.g., IL-8 in human PBMCs), and to assess its effects on cell proliferation in cell lines expressing the IL-31 receptor complex.
• In Vivo Models: It is used to induce pruritus (itch) in mice, allowing the study of scratching behavior and the mechanisms underlying itch and skin inflammation. For example, intravenous or subcutaneous injection of recombinant mouse IL-31 in mice reliably induces scratching, which is used as a model for atopic dermatitis and chronic pruritus.
• ELISA (Enzyme-Linked Immunosorbent Assay): IL-31 is used as a standard or antigen in ELISA to detect anti-IL-31 antibodies or to measure cytokine responses.
• Blocking/Neutralization Assays: Recombinant IL-31 is used to validate the efficacy of neutralizing antibodies or inhibitors targeting the IL-31 pathway, both in vitro (e.g., inhibition of IL-31-induced cell proliferation) and in vivo (e.g., reduction of IL-31-induced scratching).
• Western Blot and Immunohistochemistry: While less common, some sources indicate validation for use in Western blot and immunohistochemistry, typically for detection or quantification of IL-31 or its receptor components in tissue samples.

Supporting details and examples:

• In a published study, recombinant mouse IL-31 was used to induce scratching in BALB/c mice, and the effect was quantified over time to model pruritus.
• Functional assays include measuring the proliferation of Ba/F3 cells transfected with mouse IL-31 receptor α and OSMR genes in response to IL-31.
• Recombinant mouse IL-31 has been used to stimulate human PBMCs, resulting in IL-8 production, which is quantifiable by ELISA.
• Blocking assays have been performed using anti-IL-31 receptor antibodies to inhibit IL-31-induced biological effects, both in vitro and in vivo.
• Some commercial and research sources list validation for Western blot and immunohistochemistry, though these are less frequently described in primary research articles.

Summary of validated applications:

These applications are supported by both primary research and product validation data, with the strongest evidence for use in functional assays, in vivo pruritus models, and ELISA-based detection.

To reconstitute and prepare Recombinant Mouse IL-31 protein for cell culture experiments, follow these best-practice steps:

1. Centrifuge the vial before opening to ensure all lyophilized powder is at the bottom.

2. Reconstitution:

   • Add sterile water to achieve a final concentration of 0.1 mg/mL (100 µg/mL) unless your protocol or datasheet specifies otherwise.
   • Gently pipette the solution up and down and wash the sides of the vial to ensure complete dissolution. Do not vortex.

3. Carrier protein (optional but recommended for stability):

   • For long-term storage or if the protein will be diluted to low concentrations, add a carrier protein such as 0.1% HSA (human serum albumin) or BSA (bovine serum albumin) to prevent adsorption and loss of activity.

4. Aliquoting and storage:

   • Aliquot the reconstituted protein into small working volumes to avoid repeated freeze-thaw cycles.
   • Store aliquots at -20°C for long-term use. The reconstituted protein is typically stable for at least one week at 4°C for short-term use.

5. Dilution for cell culture:

   • Before adding to cell cultures, dilute the reconstituted stock to the desired working concentration using cell culture medium or an appropriate buffer containing carrier protein if needed.

Additional notes:

• Always refer to the lot-specific Certificate of Analysis or datasheet for any unique instructions.
• Avoid repeated freeze-thaw cycles, as this can degrade the protein.
• If using for functional assays, confirm the biological activity and endotoxin levels are suitable for your application.

Summary protocol:

1. Centrifuge vial briefly.2. Add sterile water to reach 0.1 mg/mL.3. Gently pipette to dissolve (do not vortex).4. Add 0.1% HSA or BSA if storing long-term.5. Aliquot and store at -20°C.6. Dilute to working concentration in cell culture medium before use.

These steps will ensure optimal recovery, stability, and activity of recombinant mouse IL-31 for cell culture experiments.