Recombinant Rat IL-4

Recombinant Rat IL-4 is widely used in research applications to study immune modulation, cell differentiation, tissue repair, and disease mechanisms due to its well-characterized biological activities and high purity compared to native or crude preparations.

Key scientific reasons to use recombinant rat IL-4:

• Immune Cell Differentiation and Modulation:
  IL-4 is a critical cytokine for driving the differentiation of naïve CD4+ T cells into the Th2 phenotype, which is essential for studying Th2-mediated immune responses, allergic inflammation, and asthma models. It also promotes B cell proliferation, survival, and immunoglobulin class switching, making it valuable for immunology and hematology research.

• Macrophage Polarization and Tissue Repair:
  IL-4 induces polarization of macrophages toward the M2 phenotype, which is associated with anti-inflammatory effects and tissue repair. Recombinant IL-4 has been shown to enhance tissue repair and regeneration in models of myocardial infarction and nerve injury by promoting M2 macrophage activity and stimulating fibroblast and endothelial cell function.

• Cancer and Disease Mechanisms:
  Increased IL-4 and IL-4 receptor expression are implicated in tumor progression, apoptosis resistance, and immune evasion in cancer models. Recombinant rat IL-4 enables controlled studies of these mechanisms in vitro and in vivo.

• Standardization and Reproducibility:
  Recombinant proteins offer high purity, defined activity, and batch-to-batch consistency, which are essential for reproducible results in cell culture, bioassays, and ELISA standards. This is especially important when comparing results across experiments or laboratories.

• Functional Assays:
  Recombinant rat IL-4 is used to stimulate proliferation in cell lines (e.g., TF-1 erythroleukemic cells), assess cytokine signaling pathways, and study gene expression changes in various cell types.

• Modeling Inflammatory and Autoimmune Diseases:
  IL-4 is involved in the regulation of inflammatory processes, such as those underlying rheumatoid arthritis and other autoimmune conditions, making recombinant IL-4 a valuable tool for dissecting cytokine networks and therapeutic interventions.

Typical applications include:

• Cell culture stimulation and differentiation assays
• Immune cell polarization studies
• Tissue repair and regeneration models
• ELISA standards and controls
• Mechanistic studies in cancer, allergy, and autoimmune disease models

In summary, recombinant rat IL-4 is preferred for its specificity, purity, and consistent biological activity, enabling precise investigation of cytokine-driven processes in rat-based experimental systems.

Yes, recombinant rat IL-4 can be used as a standard for quantification or calibration in ELISA assays, provided that the recombinant protein is of high purity, properly characterized, and compatible with the specific ELISA kit or assay system you are using.

Several ELISA kits for rat IL-4 (such as those from R&D Systems, Abcam, Medix Biochemica, and others) explicitly state that their assays recognize both natural and recombinant rat IL-4 and recommend using recombinant rat IL-4 as the standard for generating the calibration curve. For example:

• The R&D Systems Rat IL-4 Quantikine ELISA Kit uses E. coli-expressed recombinant rat IL-4 as the standard, and the assay is validated for accurate quantitation of both recombinant and natural rat IL-4.
• Medix Biochemica and other manufacturers also provide recombinant rat IL-4 standards with their ELISA kits, confirming that recombinant IL-4 is suitable for calibration.

Important Considerations:

• Purity and Activity: Ensure the recombinant IL-4 is highly purified and, if possible, has been validated for immunoreactivity in your specific ELISA system.
• Formulation: Some recombinant proteins may contain carrier proteins (e.g., BSA) or stabilizers that could interfere with ELISA performance. Carrier-free versions are preferred for use as standards.
• Lot-to-Lot Consistency: For reproducible results, use the same lot of recombinant IL-4 for all experiments, or re-validate the standard curve if switching lots.

In summary, recombinant rat IL-4 is commonly and appropriately used as a standard for ELISA quantification, as long as it is compatible with your assay and properly prepared.

Recombinant Rat IL-4 has been validated in published research for a range of applications, primarily focused on its immunomodulatory and cell differentiation effects in both in vitro and in vivo models.

Key validated applications include:

• Bioassays: Recombinant rat IL-4 is widely used in bioassays to study its effects on immune cell proliferation, differentiation, and function. It has been applied to:

  • Promote proliferation and survival of T cells, mast cells, basophils, eosinophils, and B cells.
  • Induce immunoglobulin class switching in B cells.
  • Drive the acquisition of the Th2 phenotype in naïve CD4+ T cells.
  • Prime and induce chemotaxis of mast cells, eosinophils, and basophils.
  • Activate and proliferate epithelial cells.

• Cell Culture: IL-4 is used to polarize macrophages to the M2 phenotype, modulate microglial activation, and study anti-inflammatory responses in various cell types, including microglia and chondrocytes.

• In Vivo Models: Recombinant rat IL-4 has been validated in animal models for:

  • Modulating neuroinflammation and improving neurological outcomes.
  • Promoting bone regeneration by regulating macrophage polarization in cranial bone defect models in rats, particularly by shifting macrophages from the pro-inflammatory M1 to the anti-inflammatory M2 phenotype.
  • Influencing tissue repair and fibrosis, as well as modulating immune responses in models of allergic inflammation, asthma, and stroke.

• Functional Assays: It has been used to induce proliferation of TF-1 human erythroleukemic cells in a dose-dependent manner, serving as a functional validation of its bioactivity.

• Immunological Studies: IL-4 is used to study cytokine signaling pathways, such as STAT6 activation, and its role in regulating the immune response, including the inhibition of NLRP3 inflammasome activation and osteoclast differentiation.

• Histological and Molecular Analyses: In vivo studies have combined IL-4 treatment with micro-CT, histomorphometry, immunohistochemistry, immunofluorescence, and RT-qPCR to assess tissue regeneration and immune modulation.

In summary, recombinant rat IL-4 is validated for use in:

• Bioassays (cell proliferation, differentiation, polarization)
• Cell culture (macrophage and microglia polarization)
• In vivo animal models (bone regeneration, neuroinflammation, tissue repair)
• Functional and molecular assays (cytokine signaling, immunomodulation).

These applications are well-supported by published research, particularly in immunology, neuroscience, and regenerative medicine contexts.

To reconstitute and prepare Recombinant Rat IL-4 protein for cell culture experiments, dissolve the lyophilized protein in sterile water or buffer to a concentration of at least 100 µg/mL, then further dilute as needed for your application. Use a carrier protein such as BSA or FCS in working solutions to stabilize the cytokine and minimize adsorption.

Step-by-step protocol:

1. Reconstitution:

   • Add sterile, endotoxin-free water or sterile PBS to the lyophilized IL-4 to achieve a concentration between 0.1–1.0 mg/mL (100–1000 µg/mL) for a stock solution.
   • If the product insert specifies, reconstitute at 50–100 µg/mL in sterile PBS containing at least 0.1% BSA or HSA (bovine or human serum albumin) to further stabilize the protein.
   • Gently mix by pipetting up and down or by slow vortexing. Avoid vigorous agitation or repeated freeze-thaw cycles, which can denature the protein.

2. Aliquoting and Storage:

   • Divide the stock solution into small aliquots to avoid repeated freeze-thaw cycles.
   • Store aliquots at ≤–20°C for long-term storage. For short-term use (up to 1 week), store at 4°C.
   • If not already present, add a carrier protein (e.g., 0.1% BSA or HSA) to aliquots for long-term stability.

3. Preparation for Cell Culture:

   • Before use, dilute the stock solution to the desired working concentration using cell culture medium or buffer containing low endotoxin and a carrier protein (e.g., 0.1% BSA or FCS).
   • Typical working concentrations for cell stimulation range from 0.1–100 ng/mL, depending on the cell type and experimental design.

4. General Best Practices:

   • Always use sterile technique to prevent contamination.
   • Centrifuge the reconstituted solution briefly to remove any insoluble material before use.
   • Avoid repeated freeze-thaw cycles by using single-use aliquots.

Summary Table:

Key points:

• Use sterile, low-endotoxin reagents throughout.
• Include a carrier protein in all solutions to stabilize IL-4.
• Store reconstituted protein in aliquots at ≤–20°C to maintain activity.

If your specific product insert provides different instructions, always follow those recommendations.