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

Recombinant Human IL-8, CXC Chemokine (77 a.a.) is widely used in research due to its well-characterized 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: IL-8 (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: IL-8 promotes tumor progression by enhancing angiogenesis, epithelial-mesenchymal 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: IL-8 directly stimulates endothelial cell proliferation and migration, supporting studies on vascular biology, wound healing, and tissue regeneration.
• Hematopoiesis and Stem Cell Studies: IL-8 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: IL-8 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 full-length, naturally occurring form of human IL-8, which is important for studies requiring physiological relevance and for comparison with post-translationally modified or truncated variants.
• Recombinant production ensures high purity and batch-to-batch 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 IL-8 (77 a.a.) provides a reliable, well-characterized tool for dissecting the molecular and cellular mechanisms underlying inflammation, immunity, cancer, and tissue repair.

Yes, you can use recombinant human IL-8 (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 IL-8 (77 a.a.) is commonly used as an ELISA standard: Many commercial ELISA kits and protocols specify the use of recombinant human IL-8, including the 77 amino acid (endothelial-derived) form, as a quantitative standard for calibration curves. This form is biologically relevant and widely accepted for quantification purposes.

• Parallelism with natural IL-8: Studies and kit documentation indicate that both the 77 a.a. and the 72 a.a. (monocyte-derived) forms of IL-8 yield parallel standard curves in validated ELISA systems, allowing accurate quantification of both recombinant and natural IL-8 in samples. This means the recombinant 77 a.a. standard is suitable for measuring IL-8 in biological matrices.

• Preparation and handling: When using recombinant IL-8 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 in-house 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 IL-8 standard used, as this can affect comparability between studies.

Summary Table: Use of Recombinant Human IL-8 (77 a.a.) as ELISA Standard

Conclusion:
Recombinant human IL-8 (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 IL-8 (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 IL-8 (77 a.a.) are standard for evaluating cell migration and receptor signaling.

• ELISA (Enzyme-Linked Immunosorbent Assay):
  Employed as a standard or control in quantifying IL-8 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, epithelial-mesenchymal transition (EMT), and immunosuppressive cell recruitment, particularly in cancer and inflammatory models.

• Western Blot:
  Used as a positive control for IL-8 detection and quantification in protein extracts.

• Blocking Assays:
  Utilized to study IL-8-mediated 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 IL-8 detection in tissue sections, aiding in localization and quantification of IL-8 expression.

Representative Research Applications:

• Cancer Biology:
  Investigating IL-8’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 siRNA-mediated IL-8 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.
• IL-8’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 phosphate-buffered 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

Short-term 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.

Long-term 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 freeze-thaw 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 pyrogen-free to prevent unwanted immune responses. Prepare aliquots of appropriate volumes (at least 10 µL per aliquot) to minimize handling and freeze-thaw 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.