Recombinant Mouse IL-5

Recombinant Mouse IL-5 is used in research applications to study and manipulate the biology of eosinophils and B cells, as it is a key cytokine regulating their differentiation, activation, proliferation, and survival.

IL-5 is a hematopoietic growth factor primarily produced by Th2 cells, mast cells, and eosinophils, and it acts through the IL-5 receptor (IL-5R). In mice, IL-5 has two major biological roles:

• Eosinophil biology: IL-5 is essential for the differentiation, maturation, activation, migration, and survival of eosinophils, making it a critical factor in models of allergic inflammation, asthma, and parasitic infection.
• B cell regulation: In mice, IL-5 also stimulates B cell growth, promotes immunoglobulin class switching (especially IgA and IgM), and supports the terminal differentiation of B cells into antibody-secreting cells.

Key research applications for recombinant Mouse IL-5 include:

• In vitro bioassays to stimulate and expand eosinophils or B cells from mouse samples.
• Cell culture supplementation to study the effects of IL-5 on immune cell differentiation, activation, and function.
• In vivo models of allergic disease, asthma, or helminth infection, where IL-5 is used to modulate eosinophil responses or to investigate the consequences of altered IL-5 signaling.
• Immunological studies on antibody production, class switching, and the interplay between T cells, B cells, and eosinophils.

Using recombinant Mouse IL-5 ensures species specificity and reproducibility, which is crucial for mechanistic studies in mouse models. It is also valuable for dissecting the molecular pathways downstream of IL-5R signaling, such as JAK–STAT and Ras/Raf-ERK pathways, which are important for immune cell survival and function.

In summary, recombinant Mouse IL-5 is a versatile tool for immunology research, particularly in studies involving eosinophil and B cell biology, allergic inflammation, and antibody responses in mouse systems.

Yes, recombinant mouse IL-5 is commonly used as a standard for quantification or calibration in ELISA assays designed to measure mouse IL-5. This is a standard practice in both commercial ELISA kits and custom assay setups.

Supporting details:

• Commercial ELISA kits for mouse IL-5 routinely use recombinant mouse IL-5 as the standard for generating the calibration curve. These kits are validated to ensure that the recombinant standard is recognized equivalently to natural mouse IL-5, allowing for accurate quantification in biological samples.
• Assay specificity and parallelism: Validation data from multiple kits show that the standard curves generated with recombinant mouse IL-5 are parallel to those obtained with natural mouse IL-5, indicating comparable antibody recognition and assay performance.
• Quantification range and sensitivity: The recombinant standard is serially diluted to generate a standard curve covering the assay’s quantification range (e.g., 15.6–1000 pg/mL or similar), and the sensitivity is typically in the low pg/mL range.
• Best practices: Each ELISA run should include a freshly prepared standard curve using the recombinant IL-5 standard to ensure accurate interpolation of sample concentrations.

Additional considerations:

• Matrix effects: Recovery experiments confirm that recombinant IL-5 can be accurately quantified in various sample matrices (e.g., serum, plasma, cell culture supernatant), supporting its use as a universal standard.
• Carrier-free vs. carrier-added: For ELISA calibration, use the recombinant IL-5 preparation recommended for ELISA (often carrier-free or with BSA, depending on kit instructions), as some preparations are optimized for bioassay rather than immunoassay applications.
• Standard preparation: Follow the specific dilution and reconstitution instructions provided with your ELISA kit or recombinant protein to ensure consistency and accuracy.

Summary:
You can confidently use recombinant mouse IL-5 as a standard for quantification or calibration in your ELISA assays, provided it is compatible with your assay’s antibodies and protocol. This approach is validated and widely accepted in research applications.

Recombinant Mouse IL-5 has been validated in published research for several key applications, primarily involving its biological activity on eosinophils and B cells, as well as its use in various in vitro and in vivo experimental systems.

Validated Applications in Published Research:

• Bioassays:
  Recombinant Mouse IL-5 is widely used in bioassays to study its effects on eosinophil differentiation, maturation, activation, migration, and survival, as well as B cell proliferation and differentiation. These assays often involve primary mouse cells or established cell lines to assess cytokine activity.

• Cell Culture:
  It is used to supplement cell culture media to promote the growth and differentiation of mouse eosinophils and B cells, and to investigate signaling pathways and gene expression regulated by IL-5.

• In Vivo Studies:
  Recombinant Mouse IL-5 has been administered to mice in preclinical models to study its physiological and pathological roles, such as in models of chronic muscle pain, allergic inflammation, asthma, and immune regulation. These studies often involve direct injection or genetic manipulation to modulate IL-5 levels in vivo.

• ELISA Standard:
  It is used as a standard in ELISA assays to quantify endogenous IL-5 levels in biological samples, supporting studies of cytokine production and immune responses.

Representative Published Research Applications:

• Eosinophil Biology:
  Used to induce eosinophil differentiation and survival in vitro and to study eosinophil-specific gene function in genetically modified mice.

• B Cell Differentiation:
  Initially identified for its ability to support the terminal differentiation of mouse B cells into antibody-secreting cells in vitro.

• Immunological Disease Models:
  Applied in models of allergic airway inflammation, food allergy, and infection to investigate the role of IL-5 in disease pathogenesis and immune regulation.

• Pain and Inflammation Research:
  Used in preclinical models to explore the role of IL-5 and regulatory T cells in chronic muscle pain and inflammatory responses.

Summary Table:

Key Notes:

• Most applications focus on the eosinophil lineage and B cell biology due to IL-5’s central role in these processes.
• Recombinant Mouse IL-5 is not intended for diagnostic or therapeutic use in humans or animals.
• The protein is validated for use in both whole cell and in vivo systems, with published studies supporting its biological activity in these contexts.

If you require protocols or more detailed application notes for a specific experimental setup, please specify the context or research focus.

To reconstitute and prepare Recombinant Mouse IL-5 protein for cell culture experiments, follow these steps for optimal solubility, stability, and biological activity:

Reconstitution Protocol:

1. Equilibrate Materials:

   • Bring the lyophilized protein vial and your chosen reconstitution buffer to room temperature before opening.

2. Centrifuge Vial:

   • Briefly centrifuge the vial to ensure all powder is at the bottom before opening.

3. Add Reconstitution Buffer:

   • Add sterile distilled water or sterile PBS (phosphate-buffered saline) to the vial. The recommended concentration for stock solutions is typically 0.1–0.5 mg/mL (100–500 μg/mL).
   • For enhanced stability and to prevent adsorption, add 0.1–1% carrier protein (such as BSA or HSA) to the buffer if the protein is carrier-free.

4. Mix Gently:

   • Allow the solution to sit at room temperature for 15–30 minutes with gentle agitation (do not vortex vigorously).
   • If visible particulates remain, continue gentle mixing for up to 2 hours at room temperature.

5. Aliquot and Storage:

   • Aliquot the reconstituted protein into sterile polypropylene tubes to avoid repeated freeze-thaw cycles.
   • Store aliquots at –20°C to –80°C for long-term use. Avoid repeated freeze-thaw cycles to preserve activity.

Preparation for Cell Culture:

• Dilution:
  Dilute the stock solution to your desired working concentration using sterile cell culture medium or buffer. For most bioassays, working concentrations range from 0.1–100 ng/mL, depending on cell type and experimental design.

• Carrier Protein:
  If the protein is carrier-free, always include 0.1% BSA or HSA in your working solutions to minimize adsorption and loss of activity.

• Sterility:
  Ensure all buffers and solutions are sterile. Filter through a 0.2 μm filter if necessary.

Additional Notes:

• Avoid harsh mixing: Do not vortex or shake vigorously, as this may denature the protein.
• Check for solubility: If the protein does not fully dissolve, gentle mixing or brief sonication may help, but avoid excessive agitation.
• Documentation: Always consult the specific product datasheet for any manufacturer-specific recommendations regarding buffer composition and concentration.

Summary Table:

These protocols ensure maximum stability and biological activity of recombinant Mouse IL-5 for cell culture experiments.