Recombinant Mouse IL-17D

Recombinant Mouse IL-17D is used in research applications to investigate its role as an immune-modulating cytokine, particularly in studies of tumor immunology, host defense against infections, and innate immune cell recruitment. It enables precise mechanistic studies by providing a controlled, consistent source of IL-17D for in vitro and in vivo experiments.

Key scientific applications and rationale include:

• Tumor Immunology: Recombinant IL-17D has demonstrated the ability to induce significant tumor growth delay and promote tumor rejection in mouse models by recruiting innate immune cells, especially natural killer (NK) cells, and stimulating chemokine production such as MCP-1 (CCL2). This makes it valuable for dissecting pathways of antitumor immunity and evaluating potential immunotherapeutic strategies.

• Innate Immune Cell Recruitment: IL-17D is a potent recruiter of innate immune cells, including neutrophils and NK cells, at sites of inflammation or infection. Its administration can be used to model early immune responses and study the dynamics of leukocyte infiltration in various tissues.

• Host Defense Against Infection: Studies show IL-17D mediates early local immune cell accumulation during viral infections (e.g., mouse cytomegalovirus), enhancing resistance to pathogens in the absence of adaptive immunity. Recombinant IL-17D is thus useful for exploring innate immune mechanisms and the interplay between cytokines and pathogen clearance.

• Cytokine Regulation and Signaling: IL-17D can stimulate the production of other cytokines (e.g., IL-6, IL-8, GM-CSF) and modulate dendritic cell activity, influencing both innate and adaptive immune responses. Recombinant protein allows for controlled studies of these signaling pathways and their downstream effects.

• Disease Models: IL-17D is implicated in inflammatory diseases, tissue repair, and immune defense. Recombinant protein is essential for modeling these processes and testing hypotheses about IL-17D’s physiological and pathological roles.

Best practices for using recombinant mouse IL-17D include:

• Dose titration to determine optimal concentrations for specific cell types or animal models.
• Use in combination with other cytokines or immune modulators to study synergistic or antagonistic effects.
• Application in both in vitro (cell culture) and in vivo (mouse models) systems for comprehensive mechanistic insights.

In summary, recombinant mouse IL-17D is a critical tool for immunology research, enabling detailed analysis of its functions in tumor rejection, immune cell recruitment, and host defense, and facilitating the development of novel therapeutic approaches.

Recombinant Mouse IL-17D can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated and matched to your assay system. Recombinant cytokines, including IL-17D, are routinely used as standards in quantitative ELISA protocols to generate standard curves for concentration determination.

Key considerations and best practices:

• Validation: Ensure the recombinant IL-17D is biologically and immunologically equivalent to the native protein detected by your assay antibodies. Most commercial ELISA kits for mouse IL-17D include recombinant IL-17D as the standard, indicating its suitability for calibration.
• Standard Curve Preparation: Prepare serial dilutions of the recombinant IL-17D in the same buffer or matrix as your samples to minimize matrix effects and ensure accurate quantification.
• Concentration Range: Use a standard curve covering the expected concentration range in your samples. Typical ELISA kits for mouse IL-17D report standard ranges from approximately 7.8–6000 pg/mL.
• Formulation: Recombinant proteins formulated with carrier proteins (e.g., BSA) are recommended for use as ELISA standards to improve stability and reproducibility.
• Documentation: Confirm the purity, concentration, and activity of your recombinant standard, ideally referencing a certificate of analysis or datasheet from the supplier.

Protocol Example:

1. Reconstitute recombinant mouse IL-17D according to the supplier’s instructions.
2. Prepare serial dilutions in assay buffer or sample matrix.
3. Run the standard curve in parallel with your samples.
4. Plot optical density (OD) versus concentration to generate the calibration curve.
5. Use the curve to interpolate sample concentrations.

Limitations:

• If your ELISA antibodies are highly specific for post-translational modifications or conformational epitopes present only in native IL-17D, confirm that the recombinant standard is recognized equivalently.
• Always verify that the recombinant standard is compatible with your specific ELISA kit or custom assay system.

In summary, recombinant mouse IL-17D is widely accepted and used as a standard for ELISA quantification, but proper validation and preparation are essential for accurate results.

Recombinant Mouse IL-17D has been validated for several important applications in published research:

Immunological Applications

Viral Infection Studies: IL-17D has been extensively studied in the context of cytomegalovirus (CMVV) infection, where it mediates early innate immune cell accumulation at sites of infection. Research demonstrates that IL-17D deficiency increases susceptibility to viral infection, establishing its role in antiviral immune responses. The protein has also been investigated for potential therapeutic applications in boosting antiviral immunity through activation of the Nrf2/IL-17D signaling axis.

Allergic Disease Research: Recombinant IL-17D has been utilized to investigate its immunoregulatory role in allergic asthma, where ciliated cell-derived IL-17D functions to restrain allergic responses.

Tumor Immunotherapy: IL-17D has been validated as a potential target for tumor immunotherapy, with studies demonstrating that injection of recombinant IL-17D into melanoma models can elicit anti-tumor immune responses.

Cytokine Production Assays

The protein stimulates production of downstream cytokines including IL-6, IL-8, and GM-CSF in endothelial cells and other cell types, making it useful for studying cytokine signaling cascades. Additionally, IL-17D inhibits hemopoiesis of myeloid progenitor cells in colony-forming assays, providing a tool for studying myeloid cell development.

Developmental Biology

Recombinant IL-17D has been validated for enhancing developmental competence in cloned embryos by inhibiting apoptosis and promoting embryonic genome activation, demonstrating applications beyond immunology in reproductive biotechnology.

Standard Laboratory Applications

For routine laboratory use, recombinant Mouse IL-17D is validated for binding assays, ELISA standards, and Western blot controls.

To reconstitute and prepare Recombinant Mouse IL-17D protein for cell culture experiments, dissolve the lyophilized protein at 100 μg/mL in sterile 4 mM HCl containing at least 0.1% human or bovine serum albumin (BSA) as a carrier protein. This approach helps maintain protein stability and prevents adsorption to surfaces.

Detailed protocol:

• Equilibrate the vial and reconstitution buffer (4 mM HCl with ≥0.1% BSA) to room temperature before opening to minimize condensation.
• Centrifuge the vial briefly to collect all lyophilized material at the bottom.
• Add the calculated volume of sterile 4 mM HCl + 0.1% BSA to achieve 100 μg/mL. For example, add 100 μL to 100 μg of protein.
• Gently mix by pipetting or slow vortexing. Avoid vigorous agitation or foaming, which can denature the protein.
• Allow to dissolve for 15–30 minutes at room temperature with gentle agitation. If undissolved material remains, continue mixing for up to 2 hours.
• Aliquot the reconstituted protein to avoid repeated freeze-thaw cycles.
• Store aliquots at –20 °C or –80 °C for long-term use. Use within two weeks after reconstitution if stored at –20 °C or –80 °C.

Additional notes for cell culture:

• Before adding to cell cultures, dilute the reconstituted stock further in cell culture medium, ideally containing 0.1% BSA or other carrier protein to maintain stability.
• Always use sterile technique to prevent contamination.
• Avoid repeated freeze-thaw cycles, as this can reduce protein activity and stability.

Summary table:

This protocol ensures optimal solubility, stability, and bioactivity of recombinant Mouse IL-17D for cell culture applications.