Recombinant Human TSLP

Recombinant Human TSLP is widely used in research applications because it is a key cytokine involved in immune regulation, particularly in the initiation and modulation of type 2 (Th2) immune responses, allergic inflammation, and hematopoietic cell differentiation.

Key scientific reasons to use recombinant human TSLP include:

• Immune Cell Activation and Differentiation: TSLP activates immature dendritic cells (DCs), promotes expansion and differentiation of B cells, and enhances proliferation of naïve and antigen-specific CD4⁺ and CD8⁺ T cells. This makes it valuable for studies on immune cell development, function, and interactions.

• Modeling Allergic and Inflammatory Diseases: TSLP is a critical mediator in asthma, atopic dermatitis, and other allergic diseases. Recombinant TSLP is used to model these conditions in vitro and in vivo, enabling investigation of disease mechanisms and therapeutic interventions.

• Functional Assays and Bioactivity Testing: Recombinant TSLP is used in bioassays to study cytokine signaling, screen for inhibitors or antibodies that block TSLP signaling, and quantify TSLP activity in various cell types.

• Therapeutic Target Validation: TSLP is a validated therapeutic target for monoclonal antibodies and small molecules in clinical development for immune-mediated diseases. Recombinant TSLP is essential for screening and characterizing these therapeutic candidates.

• Cell Culture and Differentiation Studies: TSLP supports long-term growth and differentiation of B cells, prevents apoptosis in certain cell lines, and induces chemokine release from monocytes, making it useful for cell culture optimization and mechanistic studies.

• Th2 Polarization and Cytokine Production: TSLP drives Th2 differentiation via induction of IL-4 and other Th2 cytokines, which is important for dissecting pathways of allergic inflammation and immune polarization.

Typical research applications include:

• Activation and maturation of dendritic cells for immunological studies.
• Expansion of antigen-specific T cells for adoptive cell therapy research.
• Modeling allergic inflammation and testing anti-inflammatory therapeutics.
• Investigating B cell biology and hematopoiesis.
• Screening and quantification assays for TSLP and its inhibitors.

In summary, recombinant human TSLP is a versatile tool for immunology, allergy, and inflammation research, enabling mechanistic studies, therapeutic screening, and functional assays relevant to human disease.

Yes, recombinant human TSLP can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity and properly validated for this purpose. Multiple ELISA kits and protocols specifically use recombinant human TSLP as the standard to generate calibration curves for quantifying TSLP in biological samples.

Key considerations and supporting details:

• Suitability as a Standard: Recombinant human TSLP is commonly used as a standard in commercial ELISA kits for quantifying TSLP in serum, plasma, and cell culture supernatants. These kits typically include lyophilized recombinant TSLP, which is reconstituted and serially diluted to generate a standard curve.

• Validation: The recombinant TSLP standard should be validated for use in your specific ELISA system. Commercial kits report that their recombinant TSLP standard produces standard curves that are parallel to those generated with natural TSLP, indicating comparable immunoreactivity and suitability for quantification.

• Purity and Source: Ensure the recombinant TSLP is of high purity (typically >95%) and is free from contaminants that could interfere with antibody binding or assay performance. The protein should be reconstituted and stored according to the manufacturer’s instructions to maintain stability and activity.

• Carrier Protein: Some recombinant TSLP preparations contain carrier proteins (e.g., BSA) to enhance stability. If your ELISA is sensitive to carrier proteins, use a carrier-free preparation or ensure that the carrier does not interfere with your assay.

• Bioactivity vs. Immunoreactivity: While some recombinant TSLP preparations are tested for bioactivity in cell-based assays, for ELISA calibration, the critical factor is immunoreactivity—i.e., that the standard is recognized by the capture and detection antibodies used in your assay.

• Protocol Example: Standard curves are typically prepared by serial dilution of the recombinant TSLP standard in the same buffer or matrix as your samples, and concentrations are interpolated from the standard curve.

Caveat: If you are developing a custom ELISA, confirm that your antibodies recognize the recombinant TSLP standard equivalently to native TSLP. Some antibody pairs may have different affinities for recombinant versus native forms, especially if the recombinant protein is produced in a different expression system or lacks post-translational modifications present in native TSLP.

Summary Table: Use of Recombinant Human TSLP as ELISA Standard

In conclusion, recombinant human TSLP is widely accepted and validated as a standard for ELISA quantification, provided it is compatible with your assay system and antibodies.

Recombinant Human TSLP has been validated for use in a variety of applications in published research, including:

• Bioassay: Used to stimulate cell proliferation (e.g., in BaF3 mouse pro-B cell lines transfected with human IL-7Rα and TSLPR), assess TSLP-mediated signaling, and evaluate biological activity in whole cell systems.
• ELISA: Employed as a standard or antigen for detection and quantification of TSLP.
• Functional Assay: Used to study TSLP-induced effects such as chemokine release (e.g., CCL17, CCL22), dendritic cell maturation, T cell activation, and modulation of immune responses.
• Cellular Assays: Applied in studies of TSLP signaling, screening of inhibitory molecules (e.g., anti-TSLP antibodies like tezepelumab), and reporter cell-based assays.
• Blocking Assay: Used to evaluate the efficacy of antibodies or inhibitors that block TSLP signaling.
• Western Blot: Validated for detection of TSLP protein expression.

These applications are supported by studies in primary human cells, cell lines, and animal models, with a focus on understanding TSLP's role in immune regulation, inflammation, and disease mechanisms.

To reconstitute and prepare Recombinant Human TSLP protein for cell culture experiments, follow these steps:

• Reconstitution:
  Add sterile phosphate-buffered saline (PBS) to the lyophilized protein to achieve a concentration of 100 μg/mL. If the formulation is carrier-free, use only sterile PBS; if it contains a carrier (such as BSA), use PBS with at least 0.1% human or bovine serum albumin to minimize protein loss due to adsorption.

• Mixing:
  Gently swirl or invert the vial to dissolve the protein completely. Avoid vigorous vortexing, which can denature the protein.

• Aliquoting and Storage:
  After reconstitution, aliquot the solution into small working volumes to avoid repeated freeze-thaw cycles, which can degrade the protein.

  • Store aliquots at 2–8 °C for up to one week for short-term use.
  • For long-term storage, keep aliquots at –20 °C to –70 °C.

• Preparation for Cell Culture:
  Before adding to cell cultures, dilute the reconstituted TSLP to the desired working concentration using sterile cell culture medium or PBS. Typical bioactive concentrations for cell assays range from 0.05–0.3 ng/mL for proliferation assays, but optimal concentrations should be determined empirically for your specific application.

Additional best practices:

• Centrifuge the vial briefly before opening to ensure all lyophilized material is at the bottom.
• Use sterile technique throughout to prevent contamination.
• If the protein is supplied with a specific reconstitution buffer (e.g., endotoxin-free water or a proprietary diluent), follow the manufacturer’s instructions for that lot.
• Avoid repeated freeze-thaw cycles by aliquoting immediately after reconstitution.

Summary Table: Recombinant Human TSLP Reconstitution

These guidelines ensure the stability and bioactivity of recombinant TSLP for cell culture experiments.