Recombinant Mouse B7-H2

Recombinant Mouse B7-H2 (also known as ICOSL or CD275) is a valuable tool for immunological research due to its well-characterized biological activity and versatility in experimental settings. Here are several reasons why you should consider using Recombinant Mouse B7-H2 in your research:

1. Functional Relevance in Immune Regulation

• B7-H2 is a co-stimulatory ligand that binds to ICOS (Inducible T-cell COStimulator) on activated T cells, delivering a positive signal for optimal T cell activation, proliferation, and cytokine production.
• It plays a critical role in modulating T cell responses, including the regulation of Th1, Th2, and T follicular helper (Tfh) cell differentiation.

2. Applications in Cell Culture and Functional Assays

• Recombinant Mouse B7-H2 can be used in cell culture experiments to study T cell activation, costimulation, and immune regulation.
• It is suitable for use as a positive control or stimulator in assays involving T cell proliferation, cytokine secretion, and signaling pathway analysis.

3. Standardization and Quality Control

• Recombinant Mouse B7-H2 is widely used as a standard in ELISA assays to quantify B7-H2 levels in biological samples.
• It can serve as a control in Western blotting and other immunoassays, ensuring consistency and reliability in your experimental results.

4. High Purity and Bioactivity

• Many commercially available recombinant Mouse B7-H2 proteins are produced with high purity (>95%) and low endotoxin levels, ensuring minimal interference in sensitive assays.
• The protein is often provided in lyophilized or carrier-free formats, allowing flexibility in experimental design.

5. Versatility in Research Areas

• B7-H2 is expressed by antigen-presenting cells such as B cells, macrophages, and dendritic cells, making it relevant for studies on antigen presentation, immune tolerance, and autoimmune diseases.
• It is also useful in studies of tumor immunology, transplantation, and infectious diseases where T cell costimulation is a key factor.

6. Support for Antibody and Receptor Binding Studies

• Recombinant Mouse B7-H2 can be used to study the interaction between B7-H2 and its receptors (ICOS, CD28, CTLA-4), helping to elucidate the molecular mechanisms of immune regulation.
• It is suitable for binding assays, receptor screening, and mutagenesis studies.

7. Compatibility with Multiple Assay Formats

• The protein is compatible with a wide range of applications, including ELISA, Western blot, flow cytometry, and functional cell-based assays.
• It can be immobilized on plates or used in solution, depending on the experimental requirements.

8. Research Use Only

• Recombinant Mouse B7-H2 is intended for research purposes and is not suitable for diagnostic or therapeutic use, ensuring compliance with regulatory guidelines.

In summary, Recombinant Mouse B7-H2 is a reliable, high-quality reagent that supports a broad range of immunological research applications, from basic mechanistic studies to translational research in immune-related diseases.

Yes, recombinant mouse B7-H2 is well-suited for use as a standard for quantification and calibration in ELISA assays. This application is one of the primary uses of this recombinant protein in immunoassay development.

Suitability as an ELISA Standard

Recombinant mouse B7-H2 can serve as a calibration standard because it provides a known concentration of the target protein, which is essential for generating accurate standard curves. The protein is typically available in lyophilized form and requires reconstitution according to manufacturer instructions before use.

Standard Curve Preparation

When preparing your ELISA standard curve with recombinant mouse B7-H2, follow these guidelines:

Concentration Range: Standard curves typically range from 0 to 1000 pg/mL, though this can extend to 3000 pg/mL if your predicted target protein concentration is extremely high.

Reconstitution: Most recombinant proteins are provided in lyophilized form and must be reconstituted following lot-specific instructions provided with your reagent.

Curve Quality: The R² value of your standard curve trend line should exceed 0.99 to ensure the antibody is binding properly and capturing the protein standard effectively.

Important Considerations

When using recombinant B7-H2 as your ELISA standard, ensure you are using the appropriate formulation for your application. Recombinant proteins formulated with bovine serum albumin (BSA) are generally recommended for use in cell or tissue culture and as ELISA standards. Additionally, note that ELISA standard recombinant proteins are specifically validated for calibration purposes and are not recommended for bioassay applications unless separately tested for those uses.

Recombinant Mouse B7-H2 has been validated for several key applications in published research, primarily in the context of immunology and cell signaling studies.

Validated Applications:

• Cell Culture Experiments: Recombinant Mouse B7-H2 is frequently used to study T cell activation, costimulatory signaling, and immune cell interactions in vitro. It serves as a ligand for ICOS (Inducible T-cell COStimulator) on activated T cells, enabling the investigation of costimulatory pathways and immune responses.

• ELISA (Enzyme-Linked Immunosorbent Assay): The protein is commonly used as a standard or capture reagent in ELISA assays to quantify B7-H2 or to study its interaction with ICOS and other receptors.

• Flow Cytometry: Recombinant B7-H2, or antibodies against it, are validated for flow cytometric analysis to detect expression on antigen-presenting cells or to study binding interactions with ICOS on T cells.

• Functional Assays: Published studies have used recombinant B7-H2 to assess its ability to costimulate T cell proliferation, cytokine production, and differentiation, particularly in the context of autoimmune disease models and tumor immunology. For example, administration of B7-H2 fusion proteins has been shown to potentiate T cell responses and induce tumor-specific cytotoxic T lymphocyte expansion in murine models.

• Binding Studies: Recombinant Mouse B7-H2 is validated for use in binding assays, such as surface plasmon resonance or biolayer interferometry, to characterize its interaction with ICOS and other potential receptors.

Additional Context:

• Disease Models: Recombinant B7-H2 has been used in mouse models of autoimmune diseases (e.g., lupus, arthritis) to study its role in T cell-dependent B cell responses and germinal center formation.
• Immunological Research: It is a critical reagent for dissecting the molecular mechanisms of costimulatory signaling in T cells, B cells, and dendritic cells.

Summary Table of Validated Applications

These applications are well-supported in the literature and are foundational for immunological research involving costimulatory pathways and immune cell regulation.

Reconstitution Protocol

Reconstitute lyophilized recombinant mouse B7-H2 protein by resuspending it in sterile phosphate-buffered saline (PBS) at your desired working concentration. Standard reconstitution concentrations range from 100–200 μg/mL, depending on your specific application and the product formulation. Allow the vial to reconstitute for 15–30 minutes at room temperature with gentle agitation, avoiding vigorous shaking or vortexing, which can cause foaming and protein denaturation.

Carrier Protein Considerations

When reconstituting, include a carrier protein at a final concentration of at least 0.1% to protect the recombinant protein from degradation and loss due to non-specific adsorption to container surfaces. Bovine Serum Albumin (BSA) is the most commonly used carrier protein for standard applications.

Important exception: If your experiments require serum-free culture conditions or in vivo animal studies where human or animal proteins must be excluded, use alternative stabilizers such as trehalose instead of BSA, FBS, or human serum albumin (HAS). Trehalose functions as an effective stabilizer for bioactive substances and can be used at appropriate concentrations for long-term storage.

Short-Term Storage

After reconstitution, store the protein at 2–8°C in a standard refrigerator. Under these conditions, the protein remains active for up to one week, which is suitable for typical experimental cycles of 5–7 days. For diluted working solutions, prepare them with carrier protein-containing buffer and use within one week at 4°C.

Long-Term Storage

For experiments exceeding one week or when you cannot use the entire preparation in a single experiment, dilute the reconstituted protein with a carrier protein-containing solution (0.1% BSA, 10% FBS, or 5% HAS) and store at −20°C to −80°C. For optimal stability during long-term storage, add 5–50% glycerol (final concentration) before aliquoting and freezing.

Unopened vials can be stored at room temperature or −20°C for up to 12 months when kept desiccated.

Practical Recommendations

• Do not reconstitute at protein concentrations exceeding 1 mg/mL without specific justification.
• When diluting to lower working concentrations, always use solutions containing carrier protein to maintain protein stability and bioactivity.
• Avoid repeated freeze-thaw cycles; prepare aliquots appropriate for single-use or limited reuse to preserve protein integrity.
• For cell culture applications, you may add the reconstituted protein directly to your culture medium or dilute it further depending on your experimental requirements.