Recombinant Mouse IL-3 Rα

Biological Function and Receptor Interactions

Recombinant mouse IL-3 Rα (interleukin-3 receptor alpha) serves as a critical component of the IL-3 signaling system. The IL-3 Rα subunit binds IL-3 with relatively low affinity on its own; however, when combined with the IL-3 Rβ subunit, it achieves substantially higher affinity for IL-3. This heterodimerization is essential for generating functional high-affinity receptors capable of transmitting robust growth signals.

Key Research Applications

Hematopoietic Cell Studies

Recombinant mouse IL-3 Rα is valuable for investigating hematopoietic stem cell biology and progenitor cell differentiation. IL-3 stimulates the proliferation and differentiation of hematopoietic cells across multiple lineages, including erythrocytes, megakaryocytes, granulocytes, and monocytes. This makes IL-3 Rα particularly useful for studying lineage commitment and emergency hematopoiesis, where IL-3 orchestrates extramedullary hematopoiesis in the spleen and liver during inflammatory conditions.

Immune Response Characterization

IL-3 plays a multifunctional role in immune regulation, making IL-3 Rα essential for research on innate and adaptive immunity. The cytokine enhances immune cell activation and proliferation while regulating macrophage and granulocyte production. Additionally, IL-3 modulates B cell function and influences trained immunity through monocyte activation pathways.

Inflammatory Disease Models

Recombinant mouse IL-3 Rα enables investigation of IL-3's dual roles in inflammation. In arthritis models, IL-3 inhibits TNFα-induced bone resorption and prevents cartilage degradation by reducing matrix metalloproteinase expression. This makes IL-3 Rα valuable for studying both protective and pathogenic inflammatory mechanisms across different disease contexts.

Mast Cell and Basophil Biology

IL-3 promotes mast cell growth and differentiation while enhancing basophil production during parasitic infections. Using recombinant IL-3 Rα allows researchers to examine emergency myelopoiesis and immune cell development under inflammatory conditions.

Technical Advantages

The use of recombinant IL-3 Rα provides standardized, reproducible signaling through the IL-3 receptor complex, enabling precise experimental control and quantifiable dose-response relationships in cell culture and in vivo studies.

I need to clarify an important distinction regarding your question about using recombinant mouse IL-3 Rα as an ELISA standard.

Critical Distinction: IL-3 Receptor Alpha vs. IL-3 Ligand

The recombinant protein you should use for ELISA calibration is recombinant mouse IL-3 (the cytokine ligand itself), not the IL-3 receptor alpha (IL-3Rα/CD123). These are fundamentally different molecules with distinct roles in the IL-3 signaling pathway.

Appropriate Use of Recombinant Mouse IL-3 as an ELISA Standard

Recombinant mouse IL-3 is specifically designed and validated for use as an ELISA standard. The calibration approach involves using highly purified recombinant mouse IL-3 expressed in E. coli as your reference standard. This protein typically exhibits:

• Molecular weight: Approximately 14.9 kDa as a monomer
• Purity: Greater than 98% as determined by reducing SDS-PAGE
• Specific activity: Approximately >1×10⁷ IU/mg
• Standard curve range: Typically 7.8-500 pg/mL, with sensitivity less than 10 pg/mL

Formulation Considerations

When selecting recombinant mouse IL-3 for ELISA standardization, consider the available formulations:

• With BSA carrier: Recommended for cell or tissue culture applications and as ELISA standards
• Carrier-free: Recommended when BSA presence could interfere with your specific application

Why IL-3Rα Is Not Suitable

The IL-3 receptor alpha subunit functions as a binding component of the receptor complex, not as the quantifiable analyte in sandwich ELISA assays designed to measure IL-3 cytokine levels. ELISA kits are specifically calibrated against the IL-3 ligand itself, and using the receptor protein would not provide valid quantification of IL-3 in your samples.

For accurate calibration and quantification of mouse IL-3 in your ELISA assays, use validated recombinant mouse IL-3 standards that have been characterized for this specific application.

Recombinant Mouse IL-3 Rα has been validated for several important applications in published research, though the search results primarily document applications of the full IL-3 ligand rather than the receptor alone.

Hematopoietic Cell Development and Differentiation

IL-3 and its receptor signaling have been extensively validated for stimulating hematopoietic cell proliferation and differentiation. The ligand stimulates colony formation of megakaryocytes, neutrophils, and macrophages from bone marrow cultures. Additionally, IL-3 promotes both self-renewal and differentiation of early multipotential progenitors and is involved in inducible hematopoiesis.

Bioassay Applications

Recombinant IL-3 has been validated in multiple bioassay formats, including stimulation of cell proliferation in the NFS-60 mouse myelogenous leukemia lymphoblast cell line. The receptor component has been used in functional assays to characterize IL-3 signaling responses, including STAT5 phosphorylation assays upon stimulation with recombinant IL-3.

Immune Cell Modulation

Research has demonstrated that IL-3 modulates macrophage phagocytic activity and promotes repair processes following spinal cord injury. IL-3 produced by T cells has been validated as crucial for basophil extravasation, with IL-3 inducing the expression of ALDH1A2 in primary human basophils.

Neuroinflammatory Research

IL-3 receptor signaling has been validated in studies examining microglial reprogramming in Alzheimer's disease models, where astrocyte-derived IL-3 engages IL-3Rα on microglia to ameliorate disease pathology.

Therapeutic Target Validation

The IL-3/IL-3Rα axis has been validated as a therapeutic target, with monoclonal antibodies and antibody-drug conjugates against IL-3Rα being used in clinical trials to treat hematologic cancers.

To reconstitute and prepare Recombinant Mouse IL-3 Rα protein for cell culture experiments, follow these general best practices based on standard protocols for recombinant cytokine receptor proteins:

Reconstitution Protocol

1. Centrifuge the Vial:
   Before opening, briefly centrifuge the lyophilized protein vial to ensure all powder is at the bottom.

2. Reconstitution Solution:
   Reconstitute the lyophilized protein with sterile, endotoxin-free PBS (phosphate-buffered saline) or a suitable cell culture-grade buffer (e.g., sterile water or serum-free medium).

   • Typical reconstitution concentration: 100 µg/mL (or as specified on the product datasheet).
   • Example: For a 10 µg vial, add 100 µL of buffer.

3. Gentle Mixing:
   Gently pipette the buffer down the side of the vial to dissolve the protein.

   • Do not vortex or mix vigorously to avoid denaturation.

4. Optional Carrier Protein:
   For long-term storage or to stabilize the protein, you may add a carrier protein such as 0.1% BSA (bovine serum albumin) or 5% trehalose to the reconstituted solution.

Preparation for Cell Culture

1. Aliquot and Store:

   • Aliquot the reconstituted protein into small volumes to avoid repeated freeze-thaw cycles.
   • Store aliquots at –20°C to –80°C for long-term use.

2. Working Dilution:

   • Dilute the reconstituted protein in cell culture medium or assay buffer to the desired working concentration (typically in the range of 1–100 ng/mL, depending on your experimental needs and the specific activity of the protein).

3. Sterility:

   • Ensure all steps are performed under sterile conditions if the protein will be used directly in cell culture.

Storage Recommendations

• Lyophilized: Store at –20°C to –80°C for up to 12 months.
• Reconstituted:
  • Stable at –20°C to –80°C for up to 3 months (in aliquots).
  • Can be kept at 2–8°C for up to 1 week if used immediately.

Additional Notes

• Always refer to the product datasheet or Certificate of Analysis (CoA) for specific instructions, as protocols may vary slightly between manufacturers.
• For cell culture experiments, ensure the final concentration of any carrier protein (e.g., BSA) is compatible with your cells.

Example Protocol Summary

This protocol ensures optimal activity and stability of Recombinant Mouse IL-3 Rα for cell culture applications.