Recombinant Human CXCL5/ ENA-78 (5-78 a.a.)

Recombinant Human CXCL5/ENA-78 (5-78 a.a.) is widely used in research applications due to its potent biological activities as a chemokine, particularly in studies involving neutrophil activation, chemotaxis, inflammation, angiogenesis, tissue remodeling, and cancer biology.

Key scientific reasons to use this protein in your research:

• Neutrophil Chemotaxis and Activation: CXCL5 is a strong chemoattractant and activator of neutrophils, making it essential for studying immune cell recruitment and inflammatory responses.
• Inflammation Models: CXCL5 is upregulated in response to pro-inflammatory cytokines (IL-1, TNF-α) and bacterial LPS, allowing you to model acute and chronic inflammatory conditions in vitro and in vivo.
• Angiogenesis and Tissue Remodeling: CXCL5 promotes angiogenesis and connective tissue remodeling, relevant for wound healing, fibrosis, and vascular biology studies.
• Cancer Research: CXCL5 is implicated in tumor progression, metastasis, and epithelial-mesenchymal transition (EMT), especially in colorectal and gastric cancers. Recombinant CXCL5 can be used to investigate its role in cancer cell migration, invasion, and interaction with immune cells.
• Metabolic Disease Models: CXCL5 has been shown to link obesity to insulin resistance by blocking insulin-stimulated glucose uptake in muscle, making it useful for metabolic and diabetes research.
• High Purity and Defined Activity: The recombinant 5-78 a.a. variant is a biologically active, truncated form that mimics the naturally occurring, mature chemokine, ensuring reproducible results in functional assays such as neutrophil chemotaxis and myeloperoxidase release.

Typical applications include:

• Cell migration and chemotaxis assays (e.g., Boyden chamber, transwell migration)
• Neutrophil activation studies (e.g., myeloperoxidase release, ROS production)
• Cancer cell invasion and EMT assays
• Angiogenesis assays (e.g., tube formation, endothelial cell migration)
• Inflammatory disease models (e.g., Crohn’s disease, rheumatoid arthritis)
• Metabolic and insulin signaling studies

Best practices: Use recombinant CXCL5 at concentrations validated for your specific assay (commonly 10–100 ng/mL for cell-based assays). Confirm biological activity using chemotaxis or activation assays with primary human neutrophils or relevant cell lines. Ensure endotoxin levels are low to avoid confounding inflammatory effects.

In summary, Recombinant Human CXCL5/ENA-78 (5-78 a.a.) is a versatile tool for dissecting mechanisms of immune cell recruitment, inflammation, angiogenesis, tissue remodeling, and cancer progression in controlled experimental systems.

Yes, Recombinant Human CXCL5/ENA-78 (5-78 a.a.) can be used as a standard for quantification or calibration in ELISA assays, provided it matches the form of CXCL5 detected by your assay and is validated for this use.

Key considerations and supporting details:

• Intended Use as ELISA Standard: Recombinant human CXCL5 (ENA-78, 5-78 a.a.) is specifically described as suitable for use as a standard in sandwich ELISA assays for quantifying human CXCL5. This truncated form (amino acids 5-78) is one of the naturally occurring variants and is commonly used in commercial ELISA kits.

• Validation and Compatibility: It is essential that the recombinant standard matches the epitope recognized by the capture and detection antibodies in your ELISA. Most commercial kits and protocols use recombinant CXCL5 (5-78 a.a.) as the standard, and this form is validated for accurate quantification in these systems.

• Protein Form and Activity: The 5-78 a.a. form is biologically active and structurally representative of the major circulating form of CXCL5 in humans. Ensure the recombinant protein is of high purity and properly reconstituted according to the supplier’s instructions to maintain its integrity and activity.

• Expression System: Most recombinant CXCL5 standards are produced in E. coli, which is generally acceptable for ELISA calibration, but minor differences in glycosylation or folding (compared to native protein) are unlikely to affect quantification unless your assay is sensitive to such modifications.

• Documentation: Always refer to the datasheet or technical documentation for your specific recombinant protein to confirm its suitability as an ELISA standard and to follow recommended reconstitution and storage protocols.

Summary Table: Use of Recombinant Human CXCL5 (5-78 a.a.) as ELISA Standard

Best Practices:

• Prepare a standard curve using serial dilutions of the recombinant protein in the same buffer/matrix as your samples.
• Validate the linearity and sensitivity of your assay with the chosen standard.
• Store and handle the recombinant protein according to manufacturer’s recommendations to preserve activity and prevent degradation.

If your ELISA kit or protocol specifies a different CXCL5 isoform or full-length protein, verify compatibility before use. Otherwise, recombinant human CXCL5 (5-78 a.a.) is a standard and reliable choice for ELISA calibration.

Recombinant Human CXCL5/ENA-78 (5-78 a.a.) has been validated in published research for several key applications, primarily related to its biological activity as a chemokine and its role in inflammation, immune cell recruitment, and disease models.

Validated Applications in Published Research:

• Functional Assays (Bioactivity):

  • Used to assess chemotactic activity, particularly the ability to induce migration of neutrophils and other CXCR2-expressing cells.
  • Demonstrated to stimulate myeloperoxidase release from human neutrophils, confirming its functional activity as a chemokine.
  • Applied in studies of angiogenesis, wound healing, and neovascularization using in vitro and in vivo models (e.g., endothelial cell migration, aortic ring assays, matrigel plug assays, wound healing assays).

• ELISA (Enzyme-Linked Immunosorbent Assay):

  • Used as a standard or control in ELISA to quantify CXCL5 levels in biological samples.

• Western Blot:

  • Employed as a positive control or for detection of CXCL5 in cell lysates and tissue extracts.

• Immunoprecipitation:

  • Utilized to study protein-protein interactions and signaling pathways in endothelial cells and other cell types.

• Cellular and Molecular Biology Studies:

  • Used to investigate signaling pathways (e.g., ERK/p65 activation, cytokine induction) in endothelial and immune cells.
  • Applied in studies examining insulin signaling and glucose uptake in muscle cells, linking CXCL5 to metabolic regulation.

• Disease Models:

  • Used in research on diabetes mellitus, rheumatoid arthritis, inflammatory bowel disease, and cancer to study its role in pathogenesis and as a potential therapeutic target.

Summary Table of Validated Applications

Key Notes:

• The 5-78 a.a. fragment represents the mature, biologically active form of CXCL5, commonly used in these applications.
• Most studies use concentrations in the ng/mL range for in vitro assays, with specific ED₅₀ values reported for chemotaxis and activation endpoints.
• CXCL5 is frequently used to model inflammatory responses and immune cell recruitment in both basic and translational research contexts.

If you require detailed protocols or specific references for a particular application, please specify the context or experimental system of interest.

To reconstitute and prepare Recombinant Human CXCL5/ENA-78 (5-78 a.a.) for cell culture experiments, follow these best-practice steps:

• Reconstitution:

  • Use sterile water to reconstitute the lyophilized protein.
  • A typical starting concentration is 0.1 mg/mL (100 µg/mL).
  • Gently pipette and wash down the sides of the vial to ensure full recovery of the protein into solution.
  • If higher stability is needed, you may reconstitute in sterile water or an aqueous buffer containing 0.1% BSA (bovine serum albumin) to a concentration of 0.1–1.0 mg/mL.

• Dilution for Cell Culture:

  • After reconstitution, further dilute the protein into your desired cell culture medium or buffer to the working concentration required for your experiment.
  • Typical bioactive concentrations for CXCL5 in cell-based assays range from 2–100 ng/mL, depending on the cell type and assay sensitivity.

• Handling and Storage:

  • After reconstitution, store the protein at 2–8°C for short-term use (up to 1 month).
  • For long-term storage, aliquot and freeze at –20°C to –70°C to avoid repeated freeze-thaw cycles.
  • If using BSA as a carrier, this can help stabilize the protein during storage and handling.

• General Tips:

  • Always centrifuge the vial briefly before opening to collect the lyophilized powder at the bottom.
  • Avoid vigorous vortexing, which can denature the protein; use gentle pipetting instead.
  • Ensure all solutions and materials are sterile to prevent contamination in cell culture.

Summary protocol:

1. Briefly centrifuge the vial.
2. Add sterile water (or buffer with 0.1% BSA) to achieve 0.1 mg/mL.
3. Gently pipette to dissolve.
4. Aliquot and store as needed.
5. Dilute to working concentration in cell culture medium immediately before use.

These steps will ensure optimal recovery, stability, and biological activity of recombinant CXCL5/ENA-78 for your cell culture experiments.