Recombinant Human CXCL2

Recombinant Human CXCL2 is widely used in research because it is a potent chemokine that plays a central role in neutrophil recruitment, inflammation, and tumor biology, making it valuable for studying immune cell migration, inflammatory responses, and cancer microenvironments.

Key reasons to use recombinant human CXCL2 in research applications:

• Neutrophil Chemotaxis and Activation: CXCL2 is a strong chemotactic factor for neutrophils, facilitating their recruitment and activation at sites of infection, injury, or inflammation. This makes it essential for in vitro assays investigating neutrophil migration, adhesion, and activation.
• Modeling Inflammatory Responses: CXCL2 is a critical mediator in various inflammatory diseases, including atherosclerosis, obesity, psoriasis, and rheumatoid arthritis. Recombinant CXCL2 enables controlled studies of inflammatory signaling pathways and leukocyte-endothelial interactions.
• Tumor Microenvironment Studies: CXCL2-CXCR2 signaling is implicated in tumor progression, metastasis, and immune evasion by promoting the infiltration of myeloid-derived suppressor cells and neutrophils into tumors. It is also involved in shaping the immunosuppressive microenvironment and can be used to model tumor-immune interactions or test immunotherapeutic strategies.
• Cellular and Molecular Mechanisms: Recombinant CXCL2 allows for precise dissection of downstream signaling pathways (e.g., ERK/MAPK, NF-κB, PI3K/AKT, JAK/STAT3) in target cells, supporting mechanistic studies in immunology and oncology.
• Standardization and Reproducibility: Using recombinant protein ensures batch-to-batch consistency, purity, and defined activity, which is critical for reproducible results in bioassays, cell culture, and ELISA standards.
• Versatility in Experimental Systems: Recombinant CXCL2 is suitable for a range of applications, including cell migration assays, functional studies of chemokine receptors (especially CXCR2), and as a positive control in chemotaxis or adhesion experiments.

In summary, recombinant human CXCL2 is a powerful tool for dissecting the roles of chemokines in immune cell trafficking, inflammation, and cancer, and is essential for generating reproducible, mechanistic insights in these research areas.

Yes, recombinant human CXCL2 can be used as a standard for quantification and calibration in ELISA assays. This is a well-established practice in immunoassay development and validation.

Use as a Standard in ELISA

Recombinant CXCL2 proteins serve as calibration standards for generating standard curves in sandwich ELISA formats. The recombinant protein allows you to establish a dose-response relationship between known concentrations and optical density readings, which is essential for quantifying endogenous or recombinant CXCL2 in your samples.

Key Considerations for Standard Preparation

Concentration Range and Sensitivity

Standard curves for CXCL2 ELISAs typically span ranges around 10-250 pg/mL for conventional colorimetric assays, though ultrasensitive formats can achieve detection down to 0.5 pg/mL. When preparing your standard curve with recombinant CXCL2, ensure your concentration range encompasses the expected levels in your samples.

Sample Matrix Effects

An important consideration is that your samples may contain components affecting analyte detection differently than the standard diluent provided in kits. To validate your results, prepare internal controls by spiking known amounts of recombinant CXCL2 into your actual sample matrix (serum, plasma, or cell culture supernatant). This allows you to confirm that the same recombinant CXCL2 concentration yields consistent results in both the standard diluent and your sample matrix.

Precision and Accuracy

When using recombinant CXCL2 as your standard, you can achieve excellent precision with intra-assay coefficients of variation typically below 3% and inter-assay precision around 4-6%. This demonstrates that recombinant standards provide reliable calibration for quantitative measurements.

Practical Application

Recombinant CXCL2 standards enable detection and quantification of both endogenous and recombinant CXCL2 proteins across various sample types including serum, plasma, and cell culture supernatants. Ensure you prepare a fresh standard curve for each experiment and maintain proper calibration throughout your assay runs.

Recombinant Human CXCL2 has been validated in published research primarily for bioassays measuring chemotactic activity, neutrophil recruitment, and cell signaling studies, as well as in functional studies of cancer, inflammation, and immune cell migration.

Key validated applications include:

• Bioassays for Chemotaxis and Cell Activation: Recombinant human CXCL2 is widely used to assess its ability to chemoattract cells expressing CXCR2, such as neutrophils and BaF3 mouse pro-B cells transfected with human CXCR2. These assays quantify the chemotactic potency and functional activity of CXCL2.

• Neutrophil Recruitment and Activation: Studies have used recombinant CXCL2 to demonstrate its role as a potent neutrophil attractant and activator, both in vitro and in vivo, including in models of infection, inflammation, and cancer.

• Cancer Research: Recombinant CXCL2 has been applied to investigate its effects on tumor cell migration, invasion, and metastasis, particularly in hepatocellular carcinoma and breast cancer models. It has also been used to study its role in modulating the tumor microenvironment, immune cell infiltration, and resistance to anticancer therapies.

• Inflammatory Disease Models: The protein is used to model and dissect mechanisms of chronic inflammation, such as in lung inflammation, atherosclerosis, and obesity, by driving neutrophil recruitment and activation.

• Cell Signaling Studies: Recombinant CXCL2 is employed to activate specific signaling pathways (e.g., ERK/MAPK, NF-κB, PI3K/AKT, JAK/STAT3) in target cells, allowing researchers to dissect downstream effects of CXCL2-CXCR2 axis activation.

• Hematopoietic Regulation: Certain isoforms of recombinant CXCL2 have been shown to suppress hematopoietic progenitor cell proliferation and exhibit hematoregulatory activity in vitro.

• ELISA Standard and Cell Culture Supplement: Recombinant CXCL2 is validated as a standard in ELISA assays and as a supplement in cell or tissue culture to study chemokine-mediated effects.

Additional notes:

• Most published research validates recombinant human CXCL2 in bioassays (functional chemotaxis and activation), with some studies extending to in vivo models and cell signaling analyses.
• While antibody-based applications (e.g., Western blot, ELISA, IHC) are common for CXCL2 detection, these use anti-CXCL2 antibodies rather than the recombinant protein itself.

In summary, recombinant human CXCL2 is primarily validated for functional bioassays, chemotaxis studies, immune cell recruitment, cancer and inflammation models, and as a standard in ELISA.

To reconstitute and prepare Recombinant Human CXCL2 protein for cell culture experiments, follow these steps based on best practices and manufacturer protocols:

1. Centrifuge the vial:
Briefly centrifuge the lyophilized protein vial to ensure all material is at the bottom before opening.

2. Choose the appropriate reconstitution buffer:

• For carrier-free formulations (no BSA):
  Reconstitute in sterile PBS to a concentration of 100 μg/mL.
• For formulations with BSA as a carrier:
  Reconstitute in sterile PBS containing at least 0.1% human or bovine serum albumin (BSA) to a concentration of 100 μg/mL.
• Some protocols also allow reconstitution in sterile distilled water or aqueous buffer with 0.1% BSA, especially if PBS is not suitable for your downstream application.

3. Gently mix:
Allow the protein to dissolve by gentle swirling or inversion. Avoid vigorous vortexing to prevent protein denaturation.

4. Aliquot and storage:

• Prepare small aliquots to avoid repeated freeze-thaw cycles.
• Store aliquots at –20°C to –70°C for long-term storage.
• After reconstitution, the solution is stable for up to one week at 2–8°C; for longer storage, keep at –20°C or below.

5. Working solution preparation:

• For cell culture, further dilute the stock solution to the desired working concentration using cell culture medium or appropriate buffer just before use.
• If using carrier-free protein, consider adding 0.1% BSA to the working solution to minimize adsorption to plasticware and enhance stability.

6. General notes:

• Avoid repeated freeze-thaw cycles, as this can reduce protein activity.
• Always use sterile technique to prevent contamination.

Summary Table: Reconstitution Protocol

References:

If your application is highly sensitive to carrier proteins, always use the carrier-free version and add BSA only if necessary for stability. For bioassays or cell culture, ensure the final buffer composition is compatible with your cells and assay conditions.