Recombinant Human IL-28B

Recombinant human IL-28B (also known as interferon-lambda 3 or IFN-λ3) offers several compelling advantages for research applications across virology, immunology, and therapeutic development.

Potent Antiviral Activity

IL-28B demonstrates robust antiviral efficacy against multiple viral pathogens. The protein activates the JAK-STAT signaling pathway and induces interferon-stimulated genes (ISGs), which are critical mediators of antiviral immunity. Notably, IL-28B exhibits particularly strong activity against hepatitis C virus, with studies showing IC₅₀ values as low as 0.15 × 10⁻³ mg/L in hepatocellular carcinoma cell lines. The antiviral effects can be further enhanced through combination approaches—for example, IL-28B synergistically inhibits HCV production when combined with ribavirin.

Restricted Cell-Type Tropism

Unlike broad-spectrum interferons such as IFN-α, IL-28B displays restricted cellular responsiveness, primarily affecting liver, lung, and prostate cells. This selective tropism is advantageous for research applications where you need to study tissue-specific antiviral responses without widespread systemic effects, making it ideal for investigating organ-specific immune mechanisms.

Immunomodulatory Properties

IL-28B functions as a potent immunomodulator with multiple regulatory effects. The protein suppresses Th2-mediated immune responses while promoting Th1 cytokine production, including interferon-gamma. It significantly inhibits virus-stimulated B-cell activation and antibody production—reducing B-cell proliferation and IgG secretion by over 70% in influenza-stimulated assays. Additionally, IL-28B upregulates MHC class I antigen expression, enhancing antigen presentation.

Established Production and Purification Protocols

Recombinant IL-28B can be efficiently produced using well-characterized expression systems. Methylotrophic yeast expression yields secreted protein at approximately 200 mg/L with ~95% purity achievable through simple one-step ion-exchange purification. This straightforward production methodology makes the protein accessible and cost-effective for research laboratories.

Diverse Research Applications

The protein is suitable for multiple experimental approaches, including bioassays with whole cell systems. IL-28B has demonstrated utility in studying hepatitis B and C virus replication, HIV infection of macrophages, and human metapneumovirus responses. The protein's ability to modulate both innate and adaptive immune responses makes it valuable for vaccine development research and immunological studies.

Yes, recombinant human IL-28B can be used as a standard for quantification or calibration in ELISA assays, provided it is of sufficient purity and its concentration is accurately determined. This is a common practice in quantitative ELISA protocols for cytokines and other proteins.

Essential context and best practices:

• Purity and Quantification: The recombinant IL-28B should be highly purified, ideally verified by SDS-PAGE and other analytical methods, to ensure accurate calibration. The concentration of the recombinant protein must be precisely measured, typically by absorbance at 280 nm, BCA assay, or HPLC.

• Standard Curve Preparation: Prepare a serial dilution of the recombinant IL-28B in the same buffer or diluent used for your samples to generate a standard curve. This curve allows for the quantification of IL-28B in unknown samples by comparison to the known concentrations of the standard.

• Compatibility: Most commercial ELISA kits for human IL-28B are validated to detect both natural and recombinant forms of the protein, making recombinant IL-28B suitable as a standard. Always confirm that your assay’s antibodies recognize the recombinant form you are using.

• Lot-to-Lot Variability: Be aware that different lots of recombinant protein may have slight differences in immunoreactivity or mass, which can affect quantification. It is recommended to value-assign the standard by measuring its concentration in the ELISA itself, rather than relying solely on the mass stated on the vial.

• Documentation: Follow the ELISA kit or assay protocol for standard preparation, including reconstitution instructions if the recombinant protein is lyophilized.

Additional relevant information:

• Matrix Effects: If you are quantifying IL-28B in complex biological samples (e.g., serum, plasma), ensure that your standard is prepared in a matrix that closely matches your samples to minimize matrix effects and improve accuracy.

• Validation: For publication-quality data, validate your standard curve and assay performance (e.g., linearity, sensitivity, recovery) using the recombinant IL-28B standard.

• Storage: Store recombinant IL-28B according to manufacturer or laboratory guidelines (typically at -20°C or below for long-term storage) to maintain protein integrity.

In summary, recombinant human IL-28B is scientifically appropriate for use as a standard in ELISA quantification, provided you follow best practices for protein standards and assay calibration.

Research Applications of Recombinant Human IL-28B

Recombinant human IL-28B (also known as interferon lambda 3, IFN-λ3) has been validated for several important research applications in published studies:

Antiviral Activity Studies

The primary application has been investigating antiviral mechanisms against hepatitis C virus (HCV). Recombinant IL-28B demonstrated potent anti-HCV activity in hepatoma cell cultures, with an IC₅₀ of 0.15 × 10⁻³ mg/L, and induced phosphorylation of STAT1 and expression of interferon-stimulated genes (ISGs) within one hour of treatment. The protein has also been validated in functional assays examining its ability to inhibit HCV propagation and its synergistic effects when combined with ribavirin.

Receptor and Signaling Pathway Analysis

Research has utilized recombinant IL-28B to characterize receptor usage and cellular responsiveness mechanisms. Studies have demonstrated that IL-28B signals through a heterodimeric receptor composed of IL10RB and IFNLR1, leading to JAK/STAT pathway activation. The protein has been employed in bioassays to evaluate differential expression of interferon-lambda receptor splice variants and their impact on antiviral response magnitude in human immune cells.

Cell-Type Specificity Studies

Recombinant IL-28B has been validated in bioassays examining its restricted cell tropism across different tissue types, including liver, lung, and prostate cells, demonstrating its epithelial cell-selective action compared to the broad-spectrum activity of interferon-alpha.

Viral Infection Models

The protein has been used in functional assays with various viral infection models, including vesicular stomatitis virus-infected cells and studies examining antiviral responses in human metapneumovirus and HIV infections.

To reconstitute and prepare Recombinant Human IL-28B for cell culture experiments, dissolve the lyophilized protein in sterile PBS containing at least 0.1% BSA (bovine serum albumin) to a concentration of 100 μg/mL unless otherwise specified by the product datasheet. This helps prevent protein loss due to adsorption and maintains biological activity.

Step-by-step protocol:

• Centrifuge the vial briefly before opening to ensure all lyophilized powder is at the bottom.
• Add sterile PBS with 0.1% BSA (or another recommended buffer) to achieve the desired concentration (commonly 100 μg/mL).
• Gently swirl or invert the vial to dissolve the protein. Avoid vigorous shaking or vortexing to prevent foaming and protein denaturation.
• Allow the solution to sit at room temperature for 15–30 minutes to ensure complete dissolution.
• Aliquot the reconstituted protein to avoid repeated freeze-thaw cycles, which can reduce activity.
• Storage after reconstitution:
  • Short-term (up to 1 week): Store at 2–8 °C.
  • Long-term (up to several months): Store aliquots at –20 °C to –80 °C.
• For further dilutions (e.g., to working concentrations for cell culture), use cell culture medium or PBS with 0.1% BSA to minimize protein loss.

Additional notes:

• If your experiment requires serum-free conditions, use a non-protein stabilizer such as trehalose instead of BSA.
• Always consult the specific product’s Certificate of Analysis (CoA) for any manufacturer-specific recommendations.
• Avoid repeated freeze-thaw cycles by aliquoting the protein after reconstitution.

Summary Table:

This protocol ensures maximum stability and activity of recombinant IL-28B for cell culture applications.