Recombinant Human Interleukin-33

Recombinant Human Interleukin33 (IL33) is a versatile and scientifically valuable cytokine for multiple research applications due to its diverse biological functions and wellcharacterized mechanisms of action.

Immunological Research Applications

IL33 serves as a critical regulatory cytokine with broad immunological effects. It signals through IL1RL1 and IL1RAP receptors to induce type 2 cytokine production, including IL4, IL5, and IL13, making it particularly valuable for studying allergic inflammation and immune polarization. The protein functions as a potent activator of type 2 immune responses and can be used for Th2 cell activation and maintenance studies. Additionally, IL33 contributes to host defense mechanisms by stimulating production of key inflammatory mediators.

Wound Healing and Tissue Regeneration

Recombinant IL33 demonstrates significant therapeutic potential in dermal wound healing applications. Exogenous administration of recombinant IL33 accelerates wound healing in both diabetic and nondiabetic models by increasing IL33 levels in wound tissue, elevating ILC2 cells, and promoting the transformation of macrophage M1 to M2 phenotype. At the cellular level, IL33 accelerates scratchhealing of keratinocytes and fibroblasts, providing a robust model system for studying wound repair mechanisms.

Cancer Immunotherapy Research

IL33 has emerged as a multifunctional cytokine with significant antitumor properties. The protein demonstrates synergistic interactions with various cancer therapies, including immune checkpoint blockade and chemotherapy. Recombinant IL33 enhances the antitumor activity of antiCTLA4 and antiPD1 monoclonal antibodies across different tumor models. Furthermore, IL33 plays a key role in mediating DNA damage resistance through dual mechanisms: nuclear IL33 functions as a transcriptional coactivator of homologous recombination repair genes, while the active form drives nonhomologous end joining pathways.

Intracellular Signaling Studies

IL33 is a dualfunction protein that operates both as a proinflammatory cytokine and as an intracellular nuclear factor with transcriptional regulatory capabilities. This unique characteristic makes it valuable for investigating intracellular signaling of IL1 family pathways and apoptotic/necrotic mechanisms. As a nuclearassociated cytokine normally released by damaged or necrotic cells, IL33 functions as an "alarmin"—an immediate indicator of tissue stress.

Functional Assay Applications

Recombinant IL33 is optimized for use in cell culture, differentiation studies, and functional bioassays. It has been validated in published research for cellular, molecular, and in vivo applications, including in vitro inhibition of osteoclast differentiation. The protein is suitable for ELISAbased assays and various bioassay formats with established biological activity standards.

Yes, recombinant human Interleukin33 (IL33) can be used as a standard for quantification or calibration in ELISA assays, provided it is compatible with your assay system and matches the form of IL33 detected by your antibodies.

Key considerations and supporting details:

• Recombinant IL33 is routinely used as a standard in commercial ELISA kits for generating standard curves, which are essential for quantifying IL33 concentrations in biological samples. The standard curve is constructed by measuring the optical density (OD) of known concentrations of recombinant IL33, allowing interpolation of sample concentrations.

• Form and source of recombinant IL33: Ensure that the recombinant IL33 you use as a standard matches the isoform, tag (if any), and posttranslational modifications (e.g., glycosylation) of the native protein detected in your samples. Most ELISA kits use recombinant IL33 expressed in E. coli or mammalian cells, sometimes with tags such as Histag. The standard should be as similar as possible to the endogenous IL33 in your samples for accurate quantification.

• Assay compatibility: The recombinant IL33 standard must be recognized by the capture and detection antibodies in your ELISA. Some antibodies may preferentially detect specific forms (e.g., fulllength vs. cleaved IL33, or reduced vs. oxidized forms). Check your assay’s documentation or validate the standard in your system.

• Preparation and stability: Recombinant IL33 standards are typically supplied lyophilized and should be reconstituted and used according to the manufacturer’s instructions. Many protocols recommend preparing fresh standard dilutions for each assay, as recombinant IL33 can be labile and sensitive to freeze/thaw cycles.

• Matrix effects and recovery: When spiking recombinant IL33 into biological matrices (serum, plasma, cell culture supernatant), recovery rates should be validated to ensure that the standard behaves similarly to endogenous IL33 in your assay. Some kits report recovery rates of 85–115% in various matrices, indicating good compatibility.

• Potential limitations:

  • Endogenous binding partners (e.g., soluble ST2) in samples can interfere with detection and cause underestimation of IL33 levels. Some advanced protocols include acid dissociation or blocking steps to mitigate this.
  • Lottolot variability of recombinant standards can affect quantification; using an internal calibrator or quality control sample is recommended for consistency.

Summary of best practices:

• Use recombinant human IL33 that matches the form detected by your assay.
• Prepare fresh standard dilutions for each assay run.
• Validate recovery and linearity in your sample matrix.
• Follow the ELISA kit or assay protocol for standard preparation and use.

If you provide the specific details of your ELISA system (e.g., antibody pairs, detection method), more tailored advice can be given regarding the optimal use of recombinant IL33 as a standard.

Recombinant Human Interleukin33 (IL33) has been validated in published research for a wide range of applications, including:

• Cellular and molecular studies: Used to investigate immune regulation, inflammation, tissue repair, and cancer biology.
• In vivo models: Applied in animal studies to assess immune responses, wound healing, and tumor biology.
• Wound healing: Demonstrated efficacy in promoting dermal wound healing, particularly in diabetic and nondiabetic mouse models.
• Immune cell activation: Shown to activate type 2 helper T cells (Th2), mast cells, and group 2 innate lymphoid cells (ILC2s), fostering type 2 immune responses.
• Angiogenesis and endothelial cell function: Induces urokinase and tissue factor in human endothelial cells, suggesting a role in angiogenesis and the link between coagulation and inflammation.
• Cancer biology: Enhances glioma cell migration and invasion, and activates tumor stroma to promote intestinal polyposis.
• Functional assays: Used in bioassays to study IL33 receptor expression, signaling pathways, and cytokine production.
• MicroRNA regulation: Employed to study the regulation of microRNAs involved in mast cell responsiveness and inflammation.
• Cell culture and differentiation studies: Applied in studies of cell differentiation, intracellular signaling, and inhibition of osteoclast differentiation.
• Immunotherapy: Engineered forms of IL33 have been used to expand intratumoral lymphoid cells and induce tertiary lymphoid structures for enhanced antitumor activity.

These applications highlight the versatility of recombinant human IL33 in both basic research and translational studies.

Reconstitution Protocol

Recombinant Human Interleukin33 (IL33) is supplied as a lyophilized powder and requires proper reconstitution before use in cell culture experiments. Begin by centrifuging the vial briefly to ensure all lyophilized powder settles to the bottom. Do not vortex the vial, as this can denature the protein; instead, gently pipette the reconstitution solution down the sides of the vial.

Reconstitution Solutions and Concentrations

The reconstitution medium depends on your specific application and formulation type:

• Standard formulation: Reconstitute in sterile PBS containing at least 0.1% human or bovine serum albumin (HSA or BSA) at a concentration of 10 μg/mL
• Alternative formulation: Reconstitute in sterile PBS at 100 μg/mL
• General range: Typical working concentrations range from 0.1–1.0 mg/mL (100–1000 μg/mL), with 100 μg/mL being a common stock concentration
• Highpurity water option: Reconstitute in sterile 18 MΩcm H₂O at not less than 100 μg/mL, which can then be further diluted to other aqueous solutions

Storage and Stability

Proper storage is critical for maintaining protein activity and preventing degradation.

Lyophilized protein storage:

• Store at –20°C or below in a manual defrost freezer
• Lyophilized IL33 is stable at room temperature for approximately 3 weeks but should be stored desiccated below –18°C for extended periods
• Longterm storage stability is 12 months from the date of receipt when stored at –20 to –70°C

Reconstituted protein storage:

• Shortterm storage: 2–7 days at 2–8°C (4°C) under sterile conditions
• Longterm storage: Aliquots should be stored at –20°C or below
• For extended storage beyond 3 months, add a carrier protein (0.1% HSA or BSA) to stabilize the protein
• Aliquots of reconstituted samples remain stable for up to 3 months at temperatures below –20°C

Critical Handling Considerations

Freezethaw cycles: Avoid repeated freezethaw cycles, as these significantly reduce protein activity and stability. Prepare working aliquots immediately after reconstitution to minimize handling.

Preparation technique: After reconstitution, allow the protein several minutes to fully dissolve. Do not vortex or agitate vigorously, as this can cause protein aggregation and loss of biological activity.

Sterility: Ensure all reconstitution solutions and equipment are sterile to prevent contamination and maintain protein integrity for cell culture applications.

ApplicationSpecific Considerations

Recombinant Human IL33 is optimized for cell culture, differentiation studies, and functional bioassays. The mature form of IL33 (Cterminal fragment) is the biologically active form that binds to the ST2L receptor and triggers signaling cascades involved in Th2 cell responses. When designing your experiments, ensure your reconstituted protein concentration aligns with your specific assay requirements and cell culture conditions.