Recombinant Human IL-5 Rα

Recombinant Human IL-5 Rα is a valuable tool in research applications focused on eosinophil biology, allergic inflammation, and immune signaling because it enables the study and manipulation of the IL-5 signaling pathway, which is central to eosinophil activation, differentiation, and survival.

Key reasons to use Recombinant Human IL-5 Rα in research include:

• Modeling IL-5 Signaling: IL-5 Rα is the ligand-binding subunit of the IL-5 receptor complex, which is essential for IL-5-mediated activation of eosinophils and basophils. Using recombinant IL-5 Rα allows for in vitro reconstitution of the receptor, enabling detailed studies of ligand-receptor interactions, downstream signaling, and receptor pharmacology.

• Screening Therapeutics: IL-5 Rα is a therapeutic target in diseases characterized by eosinophilia, such as asthma, hypereosinophilic syndrome, and eosinophilic granulomatosis with polyangiitis. Recombinant IL-5 Rα can be used to screen and characterize antibodies, small molecules, or biologics that block IL-5 binding or receptor activation, facilitating drug discovery and mechanistic studies.

• Studying Eosinophil Differentiation and Activation: IL-5 Rα expression is upregulated during eosinophil differentiation from hematopoietic progenitors, and its engagement by IL-5 is critical for eosinophil maturation, survival, and function. Recombinant IL-5 Rα can be used to probe these processes, including the effects of soluble versus membrane-bound receptor isoforms.

• Investigating Soluble IL-5 Rα Functions: Soluble forms of IL-5 Rα act as natural antagonists by sequestering IL-5 and preventing its interaction with cell-surface receptors, thus inhibiting downstream signaling. Recombinant soluble IL-5 Rα can be used to study this regulatory mechanism and its potential therapeutic implications.

• Assay Development: Recombinant IL-5 Rα is useful for developing and validating assays to measure IL-5 or anti-IL-5/IL-5Rα antibodies in biological samples, supporting both basic research and clinical studies.

In summary, Recombinant Human IL-5 Rα is essential for dissecting the molecular mechanisms of IL-5 signaling, modeling disease processes involving eosinophils, and supporting the development of targeted therapies for eosinophil-driven disorders.

Recombinant Human IL-5 Rα (Interleukin-5 Receptor Alpha) should not be used as a standard for quantification or calibration in ELISA assays designed to measure IL-5 cytokine levels.

Key context and supporting details:

• ELISA standards must match the analyte: In ELISA assays for IL-5 quantification, the standard must be recombinant human IL-5 itself, not its receptor (IL-5 Rα). The standard curve is generated using known concentrations of the analyte (IL-5), allowing accurate interpolation of unknown sample concentrations.

• IL-5 Rα is a different protein: IL-5 Rα is the alpha subunit of the IL-5 receptor, not the cytokine IL-5. It is structurally and functionally distinct from IL-5, and antibodies in IL-5 ELISA kits are specific for IL-5, not its receptor. Using IL-5 Rα as a standard would result in no signal or inaccurate quantification because the antibodies will not recognize or bind to the receptor in the same way as the cytokine.

• ELISA kits for IL-5 Rα exist, but are separate: There are ELISA kits specifically designed to quantify soluble IL-5 Rα (sIL-5Rα) using standards of recombinant IL-5 Rα. These are distinct from IL-5 cytokine ELISAs and require their own matched standards and antibodies.

• Best practice: Always use the exact recombinant protein that matches your target analyte as the standard for quantification in ELISA. For IL-5 quantification, use recombinant human IL-5. For IL-5 Rα quantification, use recombinant human IL-5 Rα, but only in assays specifically designed for the receptor.

Summary Table:

Conclusion:
Do not use recombinant human IL-5 Rα as a standard for IL-5 quantification ELISAs. Use only the recombinant form of the analyte you wish to measure for accurate calibration and quantification.

Recombinant Human IL-5 Rα has been validated for several key applications in published research, primarily in studies of eosinophil biology, antibody engineering, and therapeutic targeting of eosinophil-mediated diseases.

Validated Applications:

• Cellular Assays:
  Recombinant Human IL-5 Rα is widely used in in vitro assays to study eosinophil differentiation, activation, and proliferation. It serves as a target for evaluating the biological activity of anti-IL-5Rα antibodies, including inhibition of IL-5–dependent cell proliferation and induction of eosinophil apoptosis through antibody-dependent cell-mediated cytotoxicity (ADCC).

• Antibody Engineering and Epitope Mapping:
  The recombinant protein is essential for generating and characterizing monoclonal antibodies against IL-5Rα. It is used for affinity maturation, epitope mapping, and functional validation of engineered antibodies, such as those designed to block IL-5 signaling or induce eosinophil depletion.

• Flow Cytometry and Immunoassays:
  Recombinant IL-5 Rα is employed as a standard or capture antigen in flow cytometry, ELISA, and other immunoassays to quantify receptor expression or antibody binding.

• Disease Modeling and Mechanistic Studies:
  It is used in mechanistic studies to elucidate the role of IL-5Rα in eosinophilic diseases, including severe eosinophilic asthma (SEA), hypereosinophilic syndrome (HES), eosinophilic esophagitis (EE), and eosinophilic granulomatosis with polyangiitis (EGPA).

• Preclinical Therapeutic Validation:
  Recombinant IL-5 Rα is a critical reagent in preclinical models for testing the efficacy of IL-5Rα-targeting biologics, such as benralizumab and novel engineered antibodies, in reducing eosinophil counts and modulating immune responses.

Supporting Details:

• The protein is used to induce and study eosinophilic differentiation from hematopoietic progenitor cells, as well as to investigate receptor regulation and signaling pathways (e.g., PI3K, ERK, PKCδ).
• It is also applied in studies of basophil and B cell differentiation, given its expression profile.
• Recombinant IL-5 Rα is utilized in both ex vivo and in vitro settings, including assays with patient-derived eosinophils and healthy donor samples.

Summary Table:

These applications are well-supported in the literature and form the basis for ongoing research into IL-5Rα as a therapeutic target in eosinophil-associated diseases.

Reconstitution Protocol

Initial Preparation

Begin by centrifuging the lyophilized vial briefly before opening to concentrate the powder at the bottom of the tube. This step is critical to ensure accurate reconstitution and minimize protein loss.

Reconstitution Conditions

Reconstitute the protein at a concentration of 100 μg/mL in phosphate-buffered saline (PBS) containing at least 0.1% human or bovine serum albumin. When reconstituting, gently swirl or tap the vial to mix—avoid vortexing or vigorous pipetting, as mechanical stress can denature the protein and reduce its biological activity.

Storage and Stability

Short-term Storage

After reconstitution, store working aliquots at 2–8°C for up to one month. If you require longer storage, maintain the protein at –20°C to –70°C.

Long-term Storage

The lyophilized protein remains stable for 6–12 months when stored desiccated at –20°C to –70°C. Once reconstituted, the protein solution is stable at –20°C for approximately 3 months.

Critical Storage Considerations

Use a manual defrost freezer and avoid repeated freeze-thaw cycles, as these can compromise protein integrity and reduce biological activity. To minimize freeze-thaw damage, prepare aliquots of the reconstituted protein solution in appropriate volumes for your experimental needs.

Formulation Details

The lyophilized protein is typically supplied as a 0.2 μm filtered solution in PBS with bovine serum albumin as a carrier protein. The carrier protein helps stabilize the recombinant protein and prevents non-specific adsorption to container surfaces during storage and handling.