Recombinant Mouse CXCL5

Recombinant Mouse CXCL5 is a valuable tool for research applications focused on inflammation, immune cell chemotaxis, cancer biology, metabolic disease, pain mechanisms, and tissue regeneration. Its use allows precise control over experimental conditions to dissect the biological roles of CXCL5 in mouse models and cell-based assays.

Key scientific applications and rationale:

• Chemotaxis and Inflammation Studies: CXCL5 is a potent chemoattractant for neutrophils and other immune cells. Recombinant CXCL5 enables researchers to study its direct effects on immune cell migration, infiltration, and inflammatory responses in vitro and in vivo, such as in models of skin injury, gout, or infection.

• Cancer Research: CXCL5 promotes tumor cell proliferation, invasion, and metastasis, particularly via the CXCL5/CXCR2 axis. Recombinant CXCL5 is used to model its impact on tumor growth, metastatic colonization (e.g., in bone), and to test the efficacy of CXCL5/CXCR2 inhibitors.

• Metabolic Disease and Adipogenesis: CXCL5 regulates adipocyte differentiation and oxidative stress in adipose tissue. Recombinant CXCL5 can be applied to adipocyte cultures to investigate its role in lipid metabolism, insulin signaling, and ROS regulation, which is relevant for obesity and diabetes research.

• Pain and Neurobiology: CXCL5 activates nociceptive sensory neurons and contributes to pain and inflammation in models such as gouty arthritis. Recombinant CXCL5 is used to study neuronal excitability, calcium signaling, and pain pathways.

• Tissue Regeneration and Repair: CXCL5 influences epithelial regeneration, stem cell differentiation, and skin barrier restoration after injury. Recombinant CXCL5 allows mechanistic studies of its role in tissue repair and immune regulation.

Experimental advantages of recombinant protein:

• Defined concentration and purity: Recombinant CXCL5 provides a consistent and quantifiable stimulus, enabling dose-response studies and reproducible results.
• Species specificity: Mouse CXCL5 is optimal for murine models, ensuring physiological relevance and compatibility with mouse cell receptors.
• Functional assays: Recombinant CXCL5 is essential for in vitro chemotaxis, signaling, and differentiation assays, as well as for in vivo administration in mouse models.

Typical research protocols include:

• Addition of recombinant CXCL5 to cell cultures (e.g., immune cells, adipocytes, neurons) to assess migration, differentiation, or signaling responses.
• Injection or infusion in mouse models to study effects on inflammation, tumor growth, or tissue regeneration.
• Use in combination with inhibitors, neutralizing antibodies, or genetic knockouts to dissect pathway specificity and biological function.

In summary, recombinant mouse CXCL5 is indispensable for mechanistic studies of chemokine biology, immune regulation, cancer progression, metabolic disease, pain, and tissue repair in mouse systems. Its use enables precise, hypothesis-driven experimentation and translational insights into CXCL5-mediated processes.

Recombinant Mouse CXCL5 can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity, accurately quantified, and compatible with the antibodies used in your assay. Most commercial Mouse CXCL5 ELISA kits use recombinant CXCL5 as their standard for generating calibration curves.

Key considerations:

• Purity and Quantification: The recombinant CXCL5 should be highly pure (typically >95% by SDS-PAGE) and its concentration must be accurately determined, ideally by absorbance at 280 nm or amino acid analysis.
• Formulation: Standards should be reconstituted and diluted in the same buffer as recommended by your ELISA kit (often a proprietary standard diluent) to minimize matrix effects and ensure compatibility.
• Isoform and Sequence: Confirm that the recombinant CXCL5 matches the isoform and sequence recognized by the antibodies in your ELISA kit. Some kits specify the exact amino acid range used for their standard (e.g., aa 41–132).
• Validation: If using a recombinant standard not supplied with your kit, validate its performance by comparing its standard curve to the kit’s original standard. This ensures similar binding and detection characteristics.

Limitations:

• Some recombinant proteins are labeled specifically for use as ELISA standards and not for bioassays, as they may not be tested for biological activity.
• If your recombinant CXCL5 is not validated for ELISA, there may be qualitative or technical risks, such as altered epitope presentation or unexpected matrix effects.

Best Practices:

• Always run a standard curve with your recombinant CXCL5 alongside your samples.
• Use the same diluent and protocol as recommended by your ELISA kit.
• Confirm linearity, sensitivity, and reproducibility of your standard curve.

In summary, recombinant Mouse CXCL5 is suitable as an ELISA standard if it is pure, quantified, and compatible with your assay’s antibodies and protocol. Validate its performance if substituting for a kit-provided standard.

Recombinant Mouse CXCL5 has been validated in published research for several key applications, primarily in functional bioassays, cell signaling studies, and disease model investigations.

Validated Applications in Published Research:

• Bioassays:
  Recombinant Mouse CXCL5 is widely used in chemotaxis and cell migration assays, particularly to study its ability to chemoattract and activate neutrophils via the CXCR2 receptor. These assays often measure neutrophil influx or migration in response to CXCL5 stimulation.

• Adipogenesis and Metabolic Studies:
  In vitro differentiation assays using 3T3-L1 preadipocytes have shown that recombinant CXCL5 inhibits adipogenesis, as evidenced by reduced lipid droplet formation and lower triglyceride levels in treated cells. It is also used to study insulin signaling pathways, where CXCL5 enhances insulin-induced Akt phosphorylation in adipocytes.

• Cancer and Tumor Microenvironment Research:
  Recombinant CXCL5 has been used to investigate its role in tumor progression, immune cell recruitment, and metastasis. For example, it has been applied in ex vivo co-culture systems to demonstrate that CXCL5 promotes breast cancer cell proliferation and colonization in bone. It is also used in studies examining neutrophil recruitment and immune suppression in pancreatic and lung cancer models.

• Inflammation and Immune Response:
  CXCL5 is commonly used in models of pulmonary inflammation to study neutrophil migration and chemokine regulation. It is also applied in studies of wound healing and neovascularization, particularly in diabetic models, where CXCL5 neutralization or knockout is used to assess effects on vascular repair.

• Oxidative Stress and Redox Biology:
  Recombinant CXCL5 is used to assess its impact on oxidative stress and antioxidant enzyme expression in adipose tissue, especially in the context of high-fat diet-induced metabolic dysfunction.

• Pain and Sensory Biology:
  Mouse CXCL5 has been implicated in studies of pain sensitivity, particularly in models of sunburn pain, where it is used to understand chemokine-mediated modulation of nociception.

Common Experimental Techniques:

• In vitro cell culture assays (e.g., chemotaxis, differentiation, signaling)
• In vivo animal models (e.g., knockout or neutralization studies)
• Flow cytometry and immunohistochemistry (for immune cell profiling)
• Western blotting and ELISA (for signaling and cytokine quantification)
• Bioassays (for functional activity validation)

Summary Table:

These applications are supported by both direct use of recombinant protein in functional assays and by studies employing neutralization or genetic knockout to elucidate CXCL5’s biological roles.

To reconstitute and prepare Recombinant Mouse CXCL5 protein for cell culture experiments, follow these best-practice steps based on current protocols and technical recommendations:

1. Briefly centrifuge the vial
Before opening, spin the vial briefly to ensure all lyophilized protein is at the bottom.

2. Reconstitution

• Solvent: Use sterile distilled water or an aqueous buffer (such as PBS, pH 7.2–7.4) for initial reconstitution.
• Concentration: Reconstitute to a concentration between 0.1–1.0 mg/mL (100–1000 µg/mL) for a stock solution. Some protocols recommend not less than 100 µg/mL.
• Carrier protein: For enhanced stability, especially for storage or repeated freeze/thaw, add 0.1–1% BSA or HSA to the buffer.

Example calculation:
For 100 µg of protein, add 100 µL (for 1 mg/mL) up to 1 mL (for 0.1 mg/mL) of sterile water or buffer.

3. Mixing
Gently pipette up and down or swirl to dissolve. Avoid vigorous vortexing to prevent protein denaturation.

4. Aliquoting and Storage

• Aliquot the reconstituted protein into single-use volumes to avoid repeated freeze/thaw cycles.
• Short-term storage: 2–8°C for up to 1 week.
• Long-term storage: -20°C or colder (preferably -70°C) for up to several months.
• Avoid repeated freeze/thaw cycles to maintain protein activity.

5. Working solution preparation

• Dilute the stock solution to the desired working concentration using cell culture medium or appropriate assay buffer immediately before use.
• Ensure the final buffer is compatible with your cell culture system (e.g., serum-free or with low endotoxin content).

6. Endotoxin consideration

• Confirm the endotoxin level is suitable for cell culture (typically <0.1 EU/µg is acceptable for most sensitive applications).

Summary Table: Key Steps for Recombinant Mouse CXCL5 Reconstitution

Additional notes:

• Always consult the specific product datasheet for any lot-specific instructions, as formulations and recommended concentrations may vary.
• If the protein is supplied in solution (not lyophilized), it may already be in PBS and ready for dilution.

These steps will help ensure maximum activity and stability of recombinant mouse CXCL5 for your cell culture experiments.