Recombinant Human IL-8, CXC Chemokine (77 a.a.)

Recombinant Human IL8, CXC Chemokine (77 a.a.) is widely used in research due to its wellcharacterized role as a potent chemoattractant for neutrophils and its involvement in inflammation, immune regulation, angiogenesis, and cancer biology.

Key reasons to use this reagent in research applications include:

• Neutrophil Chemotaxis and Activation: IL8 (CXCL8) is the most potent human chemokine for recruiting and activating neutrophils via CXCR1 and CXCR2 receptors, making it essential for studying leukocyte migration, inflammatory responses, and innate immunity.
• Cancer Research: IL8 promotes tumor progression by enhancing angiogenesis, epithelialmesenchymal transition (EMT), immunosuppressive cell recruitment, and metastasis. It is frequently used to model tumor microenvironment interactions and to investigate mechanisms of tumor immune resistance.
• Angiogenesis and Endothelial Biology: IL8 directly stimulates endothelial cell proliferation and migration, supporting studies on vascular biology, wound healing, and tissue regeneration.
• Hematopoiesis and Stem Cell Studies: IL8 modulates myelopoiesis by suppressing myeloid progenitor proliferation and influences the homing of hematopoietic progenitors, making it useful in stem cell and bone marrow research.
• Inflammatory Disease Models: IL8 is upregulated in various inflammatory and autoimmune diseases, including atherosclerosis and rheumatoid arthritis, and is used to model these conditions in vitro and in vivo.
• Functional Assays: The recombinant 77 amino acid form is biologically active and suitable for chemotaxis assays, receptor signaling studies, and ELISA standards, ensuring reproducibility and consistency in experimental setups.

Technical advantages of the 77 a.a. form:

• The 77 a.a. isoform represents the fulllength, naturally occurring form of human IL8, which is important for studies requiring physiological relevance and for comparison with posttranslationally modified or truncated variants.
• Recombinant production ensures high purity and batchtobatch consistency, critical for quantitative and mechanistic studies.

Summary of applications:

• Leukocyte chemotaxis and migration assays
• CXCR1/CXCR2 signaling pathway studies
• Tumor microenvironment and metastasis models
• Angiogenesis and endothelial cell function assays
• Inflammatory and autoimmune disease modeling
• Hematopoietic stem cell and progenitor cell research

Using recombinant human IL8 (77 a.a.) provides a reliable, wellcharacterized tool for dissecting the molecular and cellular mechanisms underlying inflammation, immunity, cancer, and tissue repair.

Yes, you can use recombinant human IL8 (CXCL8, 77 amino acids) as a standard for quantification or calibration in ELISA assays, provided that the assay is validated for this isoform and the standard is prepared and handled according to best practices.

Key considerations and supporting details:

• Recombinant IL8 (77 a.a.) is commonly used as an ELISA standard: Many commercial ELISA kits and protocols specify the use of recombinant human IL8, including the 77 amino acid (endothelialderived) form, as a quantitative standard for calibration curves. This form is biologically relevant and widely accepted for quantification purposes.

• Parallelism with natural IL8: Studies and kit documentation indicate that both the 77 a.a. and the 72 a.a. (monocytederived) forms of IL8 yield parallel standard curves in validated ELISA systems, allowing accurate quantification of both recombinant and natural IL8 in samples. This means the recombinant 77 a.a. standard is suitable for measuring IL8 in biological matrices.

• Preparation and handling: When using recombinant IL8 as a standard, it is important to:

  • Reconstitute and dilute the protein in a buffer containing carrier protein (e.g., BSA at 5–10 mg/mL) to minimize adsorption and loss.
  • Prepare a fresh standard curve for each assay, using serial dilutions to cover the assay’s dynamic range.
  • Ensure the standard is compatible with the antibodies and detection system used in your ELISA.

• Validation: If you are using a custom or inhouse ELISA, confirm that your antibodies recognize the 77 a.a. form and that your assay demonstrates linearity, accuracy, and recovery with this standard. If using a commercial kit, refer to the kit insert to verify compatibility.

• Reporting: When publishing or reporting results, specify the isoform and source of the recombinant IL8 standard used, as this can affect comparability between studies.

Summary Table: Use of Recombinant Human IL8 (77 a.a.) as ELISA Standard

Conclusion:
Recombinant human IL8 (77 a.a.) is suitable as a standard for ELISA quantification, provided your assay is validated for this isoform and you follow best practices for standard preparation and calibration.

Recombinant Human IL8 (CXCL8, 77 a.a.) has been validated for a broad range of applications in published research, primarily due to its role as a potent neutrophil chemoattractant and mediator of inflammation and cancer biology.

Validated Applications:

• Functional/Bioassays:
  Used to assess chemotactic activity, especially neutrophil migration and activation via CXCR1 and CXCR2 receptors. Chemotaxis assays with recombinant IL8 (77 a.a.) are standard for evaluating cell migration and receptor signaling.

• ELISA (EnzymeLinked Immunosorbent Assay):
  Employed as a standard or control in quantifying IL8 levels in biological samples, including serum, plasma, and cell culture supernatants.

• Cell Culture Studies:
  Applied to stimulate cells in vitro, investigating effects on cell proliferation, survival, angiogenesis, epithelialmesenchymal transition (EMT), and immunosuppressive cell recruitment, particularly in cancer and inflammatory models.

• Western Blot:
  Used as a positive control for IL8 detection and quantification in protein extracts.

• Blocking Assays:
  Utilized to study IL8mediated signaling pathways by blocking its activity with antibodies or inhibitors, helping to dissect its role in inflammation and cancer.

• Immunohistochemistry:
  Serves as a reference or control for IL8 detection in tissue sections, aiding in localization and quantification of IL8 expression.

Representative Research Applications:

• Cancer Biology:
  Investigating IL8’s role in tumor progression, angiogenesis, metastasis, and drug resistance.
  Used in studies of glioblastoma, renal cell carcinoma, and thyroid cancer to assess its impact on cell migration, invasion, and vascularization.

• Inflammation and Immunology:
  Studying neutrophil recruitment, activation, and degranulation in response to infection or tissue injury.
  Used in models of respiratory diseases (e.g., bronchiolitis), skin ulcers, and allergic inflammation.

• Stem Cell Biology:
  Examining the role of CXCL8 in supporting pluripotency and proliferation of human pluripotent stem cells.

• Cell Survival and Rescue:
  Demonstrated to rescue normal urothelial cells from siRNAmediated IL8 knockdown, confirming its essential role in cell survival.

Summary Table of Validated Applications

Key Notes:

• The 77 amino acid form (CXCL8 6–77) is the mature, bioactive species commonly used in research.
• IL8’s validated applications span basic mechanistic studies, disease modeling, and therapeutic target validation, especially in inflammation and oncology.

If you require protocol details or specific experimental setups for any application, please specify the context.

Reconstitution Protocol

Initial Preparation

Begin by centrifuging the vial briefly to bring all lyophilized material to the bottom before opening. Avoid vortexing the sample, as this can cause protein aggregation and denaturation.

Reconstitution Solution and Concentration

Reconstitute the protein at a concentration of 0.1–1.0 mg/mL using sterile distilled water or an aqueous buffer containing 0.1% bovine serum albumin (BSA) as a carrier protein. Alternatively, you may reconstitute in sterile phosphatebuffered saline (PBS) containing at least 0.1% human or bovine serum albumin. Gently pipette the reconstitution solution down the sides of the vial and allow the protein to dissolve completely without vigorous mixing.

Storage and Stability

Shortterm Storage

Once reconstituted, the protein solution can be stored at 2–8°C for up to 1 week. For ELISA standard applications, use the reconstituted solution within 4 hours of preparation and discard any remaining material.

Longterm Storage

For extended storage beyond one week, prepare working aliquots by further diluting the protein in a buffer containing carrier protein (such as 0.1% BSA) and store at −20°C to −80°C. Use a manual defrost freezer and avoid repeated freezethaw cycles, as these can compromise protein activity and stability.

Practical Considerations for Cell Culture

When preparing the protein for cell culture experiments, ensure all solutions are sterile and pyrogenfree to prevent unwanted immune responses. Prepare aliquots of appropriate volumes (at least 10 µL per aliquot) to minimize handling and freezethaw exposure. The protein exhibits biological activity with an ED₅₀ of 0.5–2.5 ng/mL in chemoattraction assays, so appropriate dilutions should be prepared based on your specific experimental requirements.