Recombinant Human GROγ/CXCL3

Recombinant Human GROγ/CXCL3 is widely used in research due to its well-characterized roles as a chemokine involved in inflammation, immune cell recruitment, angiogenesis, and cancer biology. Its recombinant form allows for controlled, reproducible studies of its biological functions and signaling mechanisms.

Key scientific applications and rationales include:

• Chemotaxis and Immune Cell Recruitment: CXCL3 is a potent chemoattractant for neutrophils and other CXCR2-expressing cells, making it essential for studying leukocyte migration, inflammatory responses, and the mechanisms of immune cell trafficking.
• Cancer Research: CXCL3 is implicated in tumorigenesis, promoting tumor cell proliferation, migration, invasion, and angiogenesis. Exogenous recombinant CXCL3 has been shown to enhance malignant behaviors in various cancer cell lines, such as head and neck squamous cell carcinoma (HNSCC) and colon cancer, making it valuable for dissecting cancer progression and metastasis pathways.
• Signal Transduction Studies: CXCL3 activates CXCR2 (and to a lesser extent CXCR1), triggering downstream G-protein and β-arrestin signaling cascades. This makes recombinant CXCL3 a tool for investigating chemokine receptor signaling, cross-talk with other pathways (e.g., ERK1/2, STAT3, NF-κB), and pharmacological modulation.
• Inflammation and Angiogenesis Models: As a pro-inflammatory chemokine, CXCL3 is used to model acute and chronic inflammation, as well as to study its effects on endothelial cells and angiogenesis, both in vitro and in vivo.
• Biomarker and Functional Genomics Research: Elevated CXCL3 expression is associated with disease states, including cancer and inflammatory disorders. Recombinant CXCL3 enables functional genomics, transcriptomics, and biomarker validation studies.

Technical advantages of using recombinant protein:

• Defined activity and purity: Recombinant CXCL3 is produced to high purity and specific activity, ensuring reproducibility and minimizing confounding variables in experiments.
• Species specificity: Human recombinant CXCL3 is ideal for studies involving human cells or translational models, reducing cross-species variability.
• Batch-to-batch consistency: Recombinant production ensures consistent protein quality, critical for quantitative and comparative studies.

Typical research applications:

• Chemotaxis assays (e.g., transwell migration, wound healing)
• Cell proliferation and clonogenic assays
• Signal transduction and receptor binding studies
• In vivo models of inflammation, tumor growth, or metastasis
• Functional genomics and transcriptomics screens

In summary, using recombinant human GROγ/CXCL3 provides a reliable, well-characterized tool for dissecting chemokine biology, immune responses, and cancer mechanisms, supporting a wide range of experimental approaches in basic and translational research.

Yes, you can use recombinant human GROγ/CXCL3 as a standard for quantification or calibration in your ELISA assays, provided it is of high purity and its concentration is accurately known. This is a common and accepted practice in quantitative ELISA protocols for cytokines and chemokines.

Key considerations and supporting details:

• Recombinant proteins are widely used as ELISA standards: Most commercial CXCL3 ELISA kits use recombinant human CXCL3 (GROγ) as the standard to generate the calibration curve for quantification. The standard curve is typically constructed by serially diluting the recombinant protein to cover the assay’s dynamic range.

• Purity and quantification: The recombinant protein should be highly purified, and its concentration must be accurately determined (e.g., by absorbance at 280 nm, BCA assay, or HPLC). Impurities or inaccurate quantification can lead to errors in your standard curve and sample measurements.

• Form and sequence: Ensure the recombinant protein matches the native sequence and post-translational modifications relevant to your assay. Most ELISA kits use the mature form of CXCL3, and the recombinant standard should ideally match this.

• Reconstitution and handling: If your recombinant CXCL3 is lyophilized, follow the manufacturer’s or supplier’s reconstitution instructions carefully, as improper handling can affect protein stability and concentration.

• Validation: Some ELISA kits specify that their assay is validated for both natural and recombinant CXCL3, indicating that recombinant standards are suitable for calibration.

• Standard curve range: Prepare your standard curve within the range appropriate for your assay (e.g., 1.56–2000 pg/mL, depending on kit sensitivity and expected sample concentrations).

Summary table: Use of Recombinant CXCL3 as ELISA Standard

In conclusion: Using recombinant human GROγ/CXCL3 as a standard is standard practice for ELISA quantification, provided you ensure purity, accurate quantification, and compatibility with your assay system.

Recombinant Human GROγ/CXCL3 has been validated for several key applications in published research, primarily focused on its role as a chemokine in cell migration, cancer biology, and immunology.

Validated Applications:

• Cell Migration/Chemotaxis Assays:
  CXCL3 is widely used to induce chemotaxis of CXCR2-expressing cells, such as neutrophils, endothelial cells, and transfected cell lines (e.g., BaF3 mouse pro B cells expressing human CXCR2). The protein’s ability to chemoattract these cells is a standard functional validation.

• Cell Proliferation and Migration Studies in Cancer Research:
  Recombinant CXCL3 has been used to stimulate proliferation, migration, and invasion of various cancer cell lines (e.g., HSC4, KB, CAL27) in vitro. These studies typically employ proliferation assays, wound healing assays, transwell migration/invasion assays, and clonogenic assays to assess the impact of CXCL3 on tumor cell behavior.

• ELISA Standard:
  CXCL3 is validated as a standard for ELISA assays to quantify chemokine levels in biological samples. This application is essential for measuring CXCL3 concentrations in serum, plasma, or cell culture supernatants.

• Receptor Activation Studies:
  CXCL3 is used to activate CXCR2 (and sometimes CXCR1) in receptor signaling assays, including G-protein and β-arrestin pathway activation, to study downstream signaling events.

• Functional Studies in Immunology:
  CXCL3 is employed to study leukocyte migration, monocyte adhesion, and neutrophil activation, reflecting its physiological role in inflammation and immune cell trafficking.

Additional Context:

• Cancer Biology:
  CXCL3 is implicated in tumorigenesis, with research showing its involvement in tumor cell differentiation, invasion, and migration, particularly in head and neck squamous cell carcinoma (HNSCC) and other cancers.

• Inflammation Research:
  CXCL3’s role in recruiting neutrophils and basophils makes it a valuable tool for studying inflammatory responses and cytokine signaling pathways.

• Recommended Usage:
  For cell culture and ELISA, the protein is often supplied with BSA for stability, while carrier-free formulations are preferred when BSA may interfere with downstream applications.

Summary Table of Validated Applications

These applications are supported by both product validation data and peer-reviewed research, confirming the utility of recombinant human GROγ/CXCL3 in diverse experimental settings.

To reconstitute and prepare Recombinant Human GROγ/CXCL3 protein for cell culture experiments, first check the specific formulation and instructions provided with your protein, as minor differences may exist between lots or suppliers. The following protocol summarizes best practices based on multiple technical datasheets and general recombinant protein handling guidelines:

1. Reconstitution

• Centrifuge the vial briefly to collect all lyophilized powder at the bottom before opening.
• Dissolve the protein in sterile phosphate-buffered saline (PBS) or sterile water, depending on the formulation. Most sources recommend PBS, pH 7.2–7.4. If your datasheet specifies otherwise (e.g., sterile water), follow that instruction.
• Recommended concentration: Reconstitute to a stock concentration of 100–250 μg/mL. For example, add 1 mL of sterile PBS to 0.1 mg (100 μg) of protein for a 100 μg/mL solution.
• Incubate the solution at room temperature for at least 20 minutes to ensure complete dissolution.
• Do not vortex or pipette vigorously, as this may denature the protein.

2. Aliquoting and Storage

• Aliquot the reconstituted protein into small volumes to avoid repeated freeze-thaw cycles, which can reduce activity.
• Storage conditions:
  • Short-term (≤1 month): 2–8 °C under sterile conditions.
  • Long-term (up to 3 months): –20 °C to –70 °C.
  • For even longer storage, keep lyophilized at –20 °C to –70 °C, desiccated.
• Add carrier protein (e.g., 0.1% BSA) to aliquots if recommended, especially for dilute working solutions, to prevent adsorption to tube walls.

3. Preparation for Cell Culture

• Dilute the stock solution into your cell culture medium to the desired working concentration immediately before use. Typical working concentrations for chemotaxis or signaling assays are in the ng/mL range (e.g., 0.4–20 ng/mL, depending on cell type and assay).
• Filter-sterilize the final working solution if sterility is a concern and the protein is not already in a sterile buffer.

4. Additional Notes

• Check for endotoxin levels if using in sensitive cell assays; most recombinant preparations are tested to be <1 EU/μg.
• Use within one month after reconstitution for best activity.
• Avoid repeated freeze-thaw cycles to maintain protein integrity and activity.

Summary Table: Key Steps for Reconstitution and Use

Always consult the lot-specific datasheet for your recombinant protein for any unique instructions.