Recombinant Human IL-15 Rα

Using Recombinant Human IL-15 Rα in research applications is essential for maximizing the biological activity of IL-15, particularly through the mechanism of trans-presentation, which is critical for the development, survival, and activation of natural killer (NK) cells and CD8+ T cells.

Key scientific reasons to use Recombinant Human IL-15 Rα:

• Enhanced IL-15 Activity via Trans-Presentation: IL-15 alone is less effective than when complexed with IL-15 Rα. The IL-15/IL-15 Rα complex mimics physiological trans-presentation, significantly boosting the proliferation, differentiation, and survival of NK cells and CD8+ T cells compared to IL-15 alone. This is crucial for studies aiming to model or enhance immune responses.

• Improved NK and CD8+ T Cell Responses: The IL-15/IL-15 Rα complex increases the frequency and maturation of NK cells, upregulates survival proteins (such as Bcl-xL), and promotes proliferation, leading to a more robust and functional cytotoxic lymphocyte population. This is particularly valuable in immunotherapy research, cancer models, and studies of viral immunity.

• Superior In Vivo Efficacy: In animal models, administration of IL-15 complexed with IL-15 Rα (including Fc-fusion forms) results in greater expansion and activation of NK and CD8+ T cells, more effective tumor regression, and improved long-term survival compared to IL-15 alone. This makes the complex a preferred tool for preclinical studies of immune modulation.

• Modeling Human Immune Responses: Use of recombinant IL-15 Rα in humanized mouse models enables more accurate reconstitution of human NK and CD8+ T cell compartments, facilitating translational research on human immune responses to pathogens, cancer, and immunotherapies.

• Stability and Bioactivity: Recombinant complexes or fusion proteins of IL-15 and IL-15 Rα (such as those with engineered disulfide bonds or Fc domains) exhibit improved molecular stability, pharmacokinetics, and bioactivity, making them suitable for both in vitro and in vivo applications.

Typical research applications include:

• Enhancing NK and CD8+ T cell expansion in vitro for cytotoxicity assays or adoptive cell therapy protocols.
• Modeling immune reconstitution after bone marrow transplantation.
• Investigating mechanisms of cancer immunotherapy and viral immunity.
• Developing and testing IL-15-based immunotherapeutic agents.

In summary, Recombinant Human IL-15 Rα is critical for recapitulating the full biological effects of IL-15, enabling more physiologically relevant and potent immune activation in research settings.

Recombinant Human IL-15 Rα (Interleukin-15 Receptor Alpha) can only be used as a standard for quantification or calibration in ELISA assays specifically designed to detect IL-15 Rα or IL-15/IL-15Rα complexes, not for assays measuring IL-15 alone.

Key context and supporting details:

• ELISA Standard Selection: The choice of standard must match the analyte the ELISA is designed to detect. For quantifying IL-15, recombinant human IL-15 is used as the standard. For quantifying IL-15 Rα, recombinant IL-15 Rα is appropriate. For quantifying IL-15/IL-15Rα heterodimers, a recombinant heterodimer standard is required.

• Assay Specificity:

  • IL-15 ELISA: These assays use recombinant IL-15 as the standard and are validated to detect both natural and recombinant IL-15, but not IL-15 Rα alone.
  • IL-15 Rα ELISA: These assays use recombinant IL-15 Rα as the standard and are specific for the receptor, not the cytokine.
  • IL-15/IL-15Rα Complex ELISA: Specialized assays exist to detect the heterodimeric complex, requiring a standard composed of both IL-15 and IL-15Rα in a defined ratio. Using only IL-15 Rα as a standard in these assays would not yield accurate quantification of the complex.

• Scientific Best Practice: The standard must be molecularly and functionally equivalent to the analyte being measured. Using recombinant IL-15 Rα as a standard in an IL-15 ELISA would result in inaccurate calibration, as the antibodies and assay conditions are optimized for IL-15 detection, not its receptor.

• Complex Detection: If your assay is designed to detect the IL-15/IL-15Rα heterodimer, you must use a standard that is the heterodimer itself, not the individual receptor or cytokine. Studies have shown that ELISAs specific for the heterodimer do not detect single-chain IL-15 or IL-15Rα alone.

Summary Table:

Additional notes:

• Always verify the specificity of your ELISA antibodies and the intended analyte before selecting a standard.
• If your goal is to quantify IL-15 Rα or the IL-15/IL-15Rα complex, ensure your assay is validated for those targets and use the appropriate standard.

Conclusion:
Recombinant Human IL-15 Rα is only suitable as a standard for ELISA assays targeting IL-15 Rα or the IL-15/IL-15Rα complex (when complexed appropriately), but not for IL-15 quantification alone.

Recombinant Human IL-15 Rα has been validated for several key applications in published research, primarily in the context of immunology and cancer immunotherapy.

Validated Applications:

• Cellular Activation and Expansion:
  Used in vitro to activate and expand CD8+ T cells, NK cells, and NKT cells when combined with IL-15, supporting studies of immune cell proliferation and survival.

• Functional Assays:
  Applied in CTLL-2 cell proliferation assays to characterize bioactivity, especially in the context of stimulating memory CD8+ T cell expansion.

• Cancer Immunotherapy Models:
  Validated in in vivo mouse xenograft models (e.g., HT-29 xenograft in NOD-SCID mice) to assess anti-tumor efficacy via stimulation of human immune cells. Also used in combination therapies with immune checkpoint inhibitors (e.g., PD-1 antibodies) to evaluate synergistic effects on tumor growth inhibition.

• Protein-Protein Interaction and Binding Assays:
  Used in binding assays to study IL-15/IL-15Rα interactions and receptor signaling pathways.

• ELISA Standard:
  Employed as a standard in ELISA for quantifying IL-15Rα in human serum and plasma, though detection in biological samples may vary depending on assay design.

• Apoptosis Assays:
  Utilized to investigate the role of IL-15Rα in apoptosis, particularly in immune cell populations.

• Vaccine Adjuvant Studies:
  Investigated as a potential adjuvant to enhance vaccine-induced immune responses, leveraging its ability to modulate lymphocyte activity.

• Protein Characterization:
  Validated by SDS-PAGE and Western blot for identity and purity in recombinant fusion constructs (e.g., IL-15Rα-IL-15 fusion proteins).

Additional Context:

• Mechanistic Studies:
  Recombinant IL-15Rα is used to dissect IL-15/IL-2 receptor signaling, including trans-presentation mechanisms that enhance lymphocyte responses.

• Pharmacokinetic and Stability Studies:
  Fusion of IL-15Rα to IL-15 (sometimes with Fc domains) is validated for improving half-life, stability, and bioactivity in preclinical models.

• Reverse Signaling and Cellular Adhesion:
  IL-15/IL-15Rα complexes have been studied for their ability to transmit reverse signaling, affecting cellular adhesion and intracellular phosphorylation events.

These applications are supported by multiple peer-reviewed studies and are central to ongoing research in immunotherapy, immune cell biology, and translational medicine.

To reconstitute and prepare Recombinant Human IL-15 Rα protein for cell culture experiments, dissolve the lyophilized protein in sterile phosphate-buffered saline (PBS) to a concentration of 100 μg/mL, optionally including 0.1% carrier protein such as bovine serum albumin (BSA) or human serum albumin (HSA) to enhance stability and prevent adsorption to surfaces.

Detailed protocol:

• Reconstitution:

  • Briefly centrifuge the vial to collect the lyophilized powder at the bottom.
  • Add sterile PBS (pH 7.2–7.4) to achieve a final concentration of 100 μg/mL.
  • For increased stability, add 0.1% BSA or HSA to the PBS before reconstitution.
  • Gently swirl or tap the vial to mix; avoid vigorous pipetting or vortexing to prevent protein denaturation.

• Aliquoting and Storage:

  • After complete dissolution, aliquot the solution into single-use volumes to avoid repeated freeze-thaw cycles.
  • Store aliquots at –20°C or –70°C in a manual defrost freezer for long-term storage.
  • For short-term use (up to 1 month), storage at 2–8°C is acceptable.

• Working Solution Preparation:

  • Before use in cell culture, dilute the stock solution to the desired working concentration using cell culture medium or PBS containing 0.1% carrier protein.
  • Ensure all solutions are sterile and endotoxin-free, especially for sensitive cell types.

Additional notes:

• Avoid repeated freeze-thaw cycles, as this can reduce protein activity.
• If the protein is supplied as a specific domain (e.g., sushi domain), the same general reconstitution principles apply, but always consult the product datasheet for any unique requirements.
• For functional assays, confirm biological activity with a suitable bioassay after reconstitution.

This protocol is suitable for most cell culture applications, including differentiation studies and functional assays involving IL-15 signaling.