Recombinant Human Eotaxin-2

Recombinant Human Eotaxin-2 (CCL24) is a valuable tool for research applications due to its well-defined biological activity and specificity. Here are several reasons why you should consider using it in your studies:

1. Selective Chemoattractant for Eosinophils and Basophils

Eotaxin-2 is a CC chemokine that selectively attracts and activates human eosinophils and basophils via the CCR3 receptor. This makes it ideal for studies focused on allergic inflammation, asthma, and other eosinophil-driven diseases. Its specificity ensures that observed effects are directly related to eosinophil and basophil recruitment and activation, minimizing off-target effects.

2. High Purity and Bioactivity

Recombinant Eotaxin-2 is available in highly purified forms (often >95% purity by SDS-PAGE and >90% by SEC-HPLC), ensuring reliable and reproducible results in your experiments. The protein is typically lyophilized from filtered solutions and may include stabilizers like trehalose, which helps maintain its activity during storage and reconstitution.

3. Versatile Applications

• Cell Migration Assays: Eotaxin-2 can be used in chemotaxis assays to study the migration of eosinophils and basophils.
• Inflammation Studies: It is useful for investigating the mechanisms of allergic inflammation and the role of eosinophils in various disease models.
• Drug Development: Eotaxin-2 can be used to screen and evaluate potential therapeutic agents that target the eotaxin/CCR3 axis.
• ELISA and Other Immunoassays: Recombinant Eotaxin-2 can serve as a standard or positive control in ELISA and other immunoassays to quantify eotaxin-2 levels in biological samples.

4. Receptor Specificity and Cross-Desensitization

Eotaxin-2 acts exclusively via the CCR3 receptor, and its effects can be blocked by CCR3-specific antibodies. This property allows for precise manipulation and validation of CCR3-dependent pathways in your experiments. Additionally, cross-desensitization studies with eotaxin-2 and other CCR3 ligands (like eotaxin and MCP-4) can provide insights into receptor usage and signaling.

5. In Vivo and In Vitro Studies

Eotaxin-2 has been shown to induce eosinophil infiltration in vivo, making it suitable for animal models of allergic inflammation. In vitro, it can be used to study the activation and functional responses of eosinophils and basophils, such as the release of histamine and leukotriene C4.

6. Carrier-Free and BSA-Containing Options

Recombinant Eotaxin-2 is available in both carrier-free and BSA-containing forms. Carrier-free versions are ideal for cell culture and functional assays where the presence of BSA might interfere, while BSA-containing versions are suitable for use as standards in ELISA and other immunoassays.

7. Support for Advanced Research

Recombinant Eotaxin-2 is part of a broader range of cytokines and chemokines that can be used in advanced research, including organoid studies, cell therapy, and drug development. Its high quality and batch-to-batch consistency make it a reliable choice for these applications.

8. Regulatory and Clinical Relevance

The eotaxin/CCR3 axis is associated with various inflammatory and autoimmune disorders, including inflammatory bowel disease and multiple sclerosis. Using recombinant Eotaxin-2 in your research can help elucidate the role of this axis in these conditions and support the development of targeted therapies.

In summary, Recombinant Human Eotaxin-2 is a powerful and versatile tool for studying eosinophil and basophil biology, allergic inflammation, and the eotaxin/CCR3 axis. Its high purity, bioactivity, and specificity make it an excellent choice for a wide range of research applications.

Yes, recombinant human Eotaxin-2 can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity, accurately quantified, and compatible with your assay system. This is a common practice in commercial ELISA kits, where recombinant Eotaxin-2 is used to generate standard curves for quantification of the analyte in biological samples.

Key considerations and supporting details:

• Recombinant Eotaxin-2 as Standard: Multiple commercial ELISA kits for human Eotaxin-2 use recombinant protein (often expressed in E. coli) as the calibrator or standard for generating the standard curve. This approach is validated and widely accepted in quantitative immunoassays.

• Standard Curve Parallelism: Validation data from commercial kits show that standard curves generated with recombinant Eotaxin-2 are parallel to those obtained with native Eotaxin-2 in biological samples, supporting its suitability for quantification.

• Protein Quality: The recombinant protein should be of high purity, accurately quantified (e.g., by absorbance at 280 nm or amino acid analysis), and ideally carrier-free if you require precise mass-based quantification. Impurities or inaccurate concentration determination can affect the accuracy of your standard curve.

• Assay Compatibility: Ensure that the recombinant Eotaxin-2 matches the sequence and post-translational modifications (if relevant) of the target analyte in your samples. Most kits use the mature form (e.g., residues 27–119). If your ELISA antibodies are specific to certain epitopes, confirm that these are present and accessible in your recombinant standard.

• Preparation: Reconstitute and dilute the recombinant protein in the same buffer as your samples or as specified by your ELISA protocol to minimize matrix effects.

• Documentation: Some recombinant protein suppliers note that their products are suitable for use as ELISA standards, but not for bioassays unless specifically validated. Always check the product datasheet for intended use.

Summary Table: Recombinant Eotaxin-2 as ELISA Standard

In summary: Using recombinant human Eotaxin-2 as a standard for ELISA quantification is scientifically valid and widely practiced, provided the protein is of high quality and appropriately prepared for your assay system.

Recombinant Human Eotaxin-2 has been validated in published research for several key applications, primarily related to its role as a chemokine in immunology and inflammation studies.

Validated Applications in Published Research:

• In vitro chemotaxis assays: Eotaxin-2 has been used to induce chemotaxis (directed migration) of eosinophils and basophils in vitro, demonstrating its ability to selectively attract these cell types via the CCR3 receptor. This is a standard assay to study leukocyte migration and chemokine function.

• In vivo recruitment studies: Recombinant Eotaxin-2 has been injected intradermally in animal models (e.g., rhesus monkeys) to validate its ability to induce local eosinophil infiltration in tissues, confirming its chemoattractant activity in vivo.

• Histamine and leukotriene release assays: In vitro, Eotaxin-2 has been shown to stimulate histamine and leukotriene C₄ release from IL-3-primed basophils, supporting its role in allergic inflammation.

• Calcium flux assays: Eotaxin-2 has been used to induce intracellular calcium mobilization in eosinophils, further validating its activity through CCR3 signaling pathways.

• Colony formation suppression: It has been reported as a strong suppressor of colony formation by multipotential hematopoietic progenitor cell lines, indicating a role in hematopoietic regulation.

• Cytokine profiling and biomarker studies: Eotaxin-2 levels have been measured in serum or tissue samples as part of multiplex cytokine profiling, particularly in studies of allergic diseases (e.g., allergic rhinitis, asthma) to assess its value as a biomarker for disease activity or therapeutic response.

• ELISA standard: Recombinant Eotaxin-2 is widely used as a standard in ELISA assays for the quantitative measurement of endogenous Eotaxin-2 in biological samples.

Summary Table: Applications of Recombinant Human Eotaxin-2

Key Context:

• Eotaxin-2 is highly selective for CCR3-expressing cells, making it a valuable tool for dissecting eosinophil and basophil biology in allergy and inflammation research.
• Its use as an ELISA standard and in cytokine profiling supports translational and clinical research into allergic and autoimmune diseases.

If you require protocols or more specific experimental details for any of these applications, please specify the intended use.

To reconstitute and prepare Recombinant Human Eotaxin-2 (CCL24) for cell culture experiments, briefly centrifuge the vial to collect the lyophilized powder, then reconstitute in sterile distilled water or aqueous buffer containing 0.1% BSA to a concentration between 0.1–1.0 mg/mL. After reconstitution, gently mix the solution to ensure complete dissolution.

Detailed protocol and best practices:

• Centrifugation: Before opening, briefly centrifuge the vial to ensure all powder is at the bottom.
• Reconstitution buffer: Use sterile distilled water or a buffer (e.g., PBS) containing 0.1% BSA as a carrier protein to stabilize the protein and prevent adsorption to surfaces.
• Concentration: Aim for a final concentration of 0.1–1.0 mg/mL. For example, add 100–1000 µL of buffer to 100 µg of protein.
• Mixing: Gently mix (do not vortex) until the protein is fully dissolved.
• Aliquoting: Divide the stock solution into small aliquots to avoid repeated freeze-thaw cycles.
• Storage: Store aliquots at –20°C or colder for long-term use. For short-term use (up to 1 month), aliquots can be kept at 2–8°C.
• Avoid freeze-thaw cycles: Repeated freeze-thawing can denature the protein and reduce activity.
• Working solution: Dilute the stock solution to the desired working concentration in cell culture medium immediately before use. Always use sterile technique.

Additional notes:

• If the protein is to be used in sensitive cell culture assays, confirm that the endotoxin level is suitable (typically <1 EU/µg).
• Carrier protein (BSA) is recommended for stability, especially at low concentrations or during storage.
• After reconstitution, the protein is generally stable for several months at –20°C in aliquots with carrier protein.

Summary Table: Preparation Steps

This protocol ensures optimal stability and biological activity of recombinant Eotaxin-2 for cell culture applications.