Recombinant Human CXCL5

Using Recombinant Human CXCL5 in research applications allows you to directly investigate the biological functions and signaling mechanisms of this chemokine in controlled experimental systems. Recombinant CXCL5 is particularly valuable for dissecting its roles in cell migration, inflammation, adipogenesis, oxidative stress regulation, and cancer biology.

Key scientific applications and rationale include:

• Functional Studies: Recombinant CXCL5 enables precise analysis of its effects on target cells, such as inducing chemotaxis in neutrophils or CXCR2-expressing cells, and modulating immune cell migration in vitro bioassays.
• Adipogenesis and Metabolism: CXCL5 has been shown to inhibit adipocyte differentiation and reduce triglyceride accumulation in preadipocyte models. Treatment with recombinant CXCL5 during adipogenesis decreases lipid droplet formation and downregulates adipogenic markers, making it a tool for studying metabolic regulation and fat cell biology.
• Oxidative Stress and Inflammation: CXCL5 modulates reactive oxygen species (ROS) production and antioxidant protein expression. Recombinant CXCL5 can be used to probe its impact on oxidative stress pathways and inflammatory responses in various cell types.
• Cancer Research: CXCL5 is implicated in tumor progression, metastasis, and angiogenesis. Recombinant CXCL5 has been demonstrated to promote cancer cell proliferation and colonization, particularly in the context of the bone microenvironment and through the CXCL5/CXCR2 axis. This makes it essential for modeling tumor–microenvironment interactions and testing therapeutic inhibitors.
• Receptor-Ligand Studies: Using recombinant CXCL5 allows for controlled activation of its receptor, CXCR2, facilitating studies on downstream signaling, receptor pharmacology, and antagonist screening.
• Standardization and Reproducibility: Recombinant proteins provide a consistent, defined reagent for quantitative assays (e.g., ELISA standards, dose-response studies), improving reproducibility across experiments.

In summary, recombinant human CXCL5 is a critical reagent for mechanistic studies in immunology, metabolism, and oncology, enabling controlled, hypothesis-driven experiments that elucidate its diverse biological roles.

Yes, recombinant human CXCL5 can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated and matched to the assay system. Recombinant CXCL5 is commonly used as a standard in sandwich ELISA formats to generate a calibration curve for quantifying CXCL5 concentrations in biological samples.

Key considerations and best practices:

• Assay Compatibility: Ensure the recombinant CXCL5 standard is compatible with the antibodies used in your ELISA. Most commercial ELISA kits for CXCL5 are validated to recognize both natural and recombinant forms of the protein, and their standard curves are typically generated using recombinant CXCL5.
• Standard Preparation: Follow the manufacturer’s instructions for reconstitution and dilution of the recombinant standard. Accurate preparation is critical for reliable quantification.
• Validation: Confirm that the recombinant standard produces a dose-response curve parallel to that of endogenous CXCL5 in your sample matrix. This ensures that quantification is accurate and not affected by matrix effects or differences between recombinant and native protein.
• Purity and Formulation: Use recombinant CXCL5 of high purity and appropriate formulation (e.g., with or without carrier proteins like BSA) as recommended for ELISA standards.
• Documentation: Record lot numbers and concentrations for reproducibility and traceability in your experiments.

Scientific applications:

• Recombinant CXCL5 standards are used to quantify CXCL5 in serum, plasma, and cell culture supernatants in research on inflammation, cancer, and immune responses.
• Calibration with recombinant standards allows for precise measurement of CXCL5 levels, facilitating comparison across experiments and laboratories.

Protocol summary:

1. Reconstitute recombinant CXCL5 according to instructions.
2. Prepare serial dilutions to generate a standard curve covering the expected concentration range.
3. Run standards alongside samples in the ELISA.
4. Use the standard curve to interpolate CXCL5 concentrations in unknown samples.

Conclusion:
Using recombinant human CXCL5 as a standard is a scientifically accepted and validated approach for ELISA quantification, provided assay-specific validation steps are followed.

Recombinant Human CXCL5 has been validated for several key applications in published research, primarily in the fields of immunology, cell biology, cancer biology, and vascular biology.

Validated Applications:

• Bioassays (Chemotaxis and Cell Migration):

  • Recombinant human CXCL5 is widely used in in vitro chemotaxis assays to assess its ability to attract cells expressing CXCR2, such as neutrophils and engineered cell lines (e.g., Baf3-mCXCR2 transfectants). Dose-dependent chemotactic activity has been demonstrated, with effective concentrations (ED50) typically in the range of 10–60 ng/mL.
  • It is also used to study migration and network formation of endothelial cells and endothelial progenitor cells, particularly in the context of angiogenesis and wound healing.

• Cell Proliferation and Functional Assays:

  • Recombinant CXCL5 has been shown to promote proliferation of cancer cells, such as papillary thyroid carcinoma (PTC) cells, in functional cell-based assays.
  • It is used to study the activation of signaling pathways (e.g., ERK/p65) and the regulation of inflammatory cytokines (IL-1β, IL-6, TNF-α) in various cell types.

• Protein-Protein Interaction Studies:

  • CXCL5 is employed in protein-protein interaction analyses, particularly to investigate its binding to the CXCR2 receptor and downstream signaling events.

• Immunological Research:

  • Used to model neutrophil recruitment and activation in inflammation and cancer microenvironments, including studies of immune cell infiltration in tumor models.

• Angiogenesis and Neovascularization Assays:

  • Recombinant CXCL5 is validated in in vitro and in vivo angiogenesis assays, such as tube formation, aortic ring, and matrigel plug assays, to study its pro-angiogenic effects.

• ELISA and Immunoassay Standards:

  • It is used as a standard in immunoassays (e.g., ELISA) for quantifying CXCL5 levels in biological samples.

• SDS-PAGE and Quality Control:

  • The protein is routinely validated by SDS-PAGE for purity and molecular weight confirmation.

Summary Table of Validated Applications

Additional Notes:

• Most studies use recombinant CXCL5 at concentrations ranging from 10 ng/mL to 10 µg/mL, depending on the assay and cell type.
• Functional validation is typically performed in human and mouse cell systems, with cross-species activity often reported.
• The protein is also used in therapeutic development research, particularly as a target for modulating inflammation, angiogenesis, and tumor progression.

If you require protocol-specific details or validation in a particular model system, please specify the context for more targeted information.

To reconstitute and prepare Recombinant Human CXCL5 protein for cell culture experiments, dissolve the lyophilized protein in sterile water to a concentration of 0.1 mg/mL (100 μg/mL) or higher, then gently mix to ensure complete solubilization. The resulting stock can be further diluted in appropriate sterile buffers or cell culture media for experimental use.

Detailed protocol and best practices:

• Centrifuge the vial briefly before opening to collect all lyophilized material at the bottom.
• Add sterile water (or sterile PBS if specified by your protocol) to achieve a concentration of at least 100 μg/mL. For example, add 100 μL of water to 10 μg of protein.
• Gently pipet up and down to dissolve the protein, washing down the sides of the vial to recover all material.
• Avoid vigorous vortexing to minimize protein denaturation.
• Incubate at room temperature for several minutes if necessary to ensure full dissolution.
• Aliquot the stock solution to avoid repeated freeze-thaw cycles, which can degrade the protein.
• Store aliquots at −20 °C or below for long-term storage; short-term storage at 4 °C is acceptable for up to one month.
• Dilute the stock into cell culture medium or buffer immediately before use to the desired working concentration. If using serum-free media, consider adding carrier proteins (e.g., 0.1% BSA) to minimize adsorption to plasticware.

Additional considerations:

• Endotoxin levels: Ensure the recombinant CXCL5 preparation is endotoxin-free (<0.01 EU/μg) for sensitive cell culture applications.
• Sterility: Use sterile technique throughout to prevent contamination.
• Avoid repeated freeze-thaw cycles by aliquoting the stock solution.
• Working concentrations: Typical experimental concentrations for chemokines like CXCL5 range from 1–100 ng/mL, but optimal dosing should be determined empirically for your cell type and assay.

Summary Table:

These steps will ensure that your recombinant CXCL5 is properly prepared for reliable and reproducible cell culture experiments.