Recombinant Mouse CD120a (TNFR1)

Recombinant Mouse CD120a (TNFR1) is a valuable tool for research applications focused on immune regulation, inflammation, apoptosis, and tumor biology because it is the principal receptor for TNF-α, mediating key cellular responses through its death domain and high-affinity ligand binding.

Key scientific applications and rationale:

• Mechanistic Studies of TNF Signaling: TNFR1 (CD120a) is the main receptor for soluble TNF-α, initiating downstream signaling pathways that regulate apoptosis, cell survival, and inflammation. Recombinant TNFR1 allows for controlled in vitro studies of ligand-receptor interactions, receptor activation, and downstream signaling events.

• Immune Modulation and Disease Models: TNFR1 is central to immune cell regulation. Its signaling can promote or inhibit immune responses, as shown in models of cancer and autoimmune disease. Recombinant TNFR1 can be used to dissect these pathways, test antagonists or agonists, and study immune cell behavior in response to TNF-α.

• Apoptosis and Cell Death Research: The death domain of TNFR1 interacts with adaptor proteins to trigger apoptosis. Recombinant TNFR1 enables detailed biochemical and cellular assays to study programmed cell death mechanisms.

• Tumor Biology and Immunotherapy: TNFR1 signaling has been implicated in tumor progression and immune evasion. For example, TNFR1 blockade increases dendritic cell infiltration and anti-tumor immunity in mouse models of pancreatic cancer. Recombinant TNFR1 can be used to screen for inhibitors, study tumor microenvironment interactions, and develop therapeutic strategies.

• Protein-Protein Interaction Assays: Recombinant TNFR1 is essential for binding studies, such as surface plasmon resonance, ELISA, or co-immunoprecipitation, to characterize TNF-α or drug candidates targeting TNFR1.

• Structural and Biophysical Analysis: Recombinant forms facilitate crystallography, NMR, or other structural studies to elucidate receptor conformation, ligand binding sites, and domain organization.

Best practices for use:

• Use recombinant TNFR1 in in vitro assays to precisely control receptor concentration and avoid confounding effects from endogenous expression.
• Employ recombinant protein in cell-based assays to study receptor activation, signaling, and functional outcomes.
• Validate recombinant TNFR1 activity and purity before use in sensitive applications such as apoptosis or immune cell assays.

Summary of advantages:

• High specificity and reproducibility for mechanistic studies.
• Enables quantitative and structural analyses not possible with native receptor alone.
• Facilitates drug screening and antibody validation for immunomodulatory therapies.

Recombinant Mouse CD120a (TNFR1) is thus indispensable for dissecting TNF-α biology, modeling disease mechanisms, and developing targeted interventions in immunology, oncology, and cell death research.

Yes, recombinant Mouse CD120a (TNFR1) protein can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated for this purpose. Recombinant TNFR1 is commonly used as a standard in commercial mouse TNFR1 ELISA kits, which are designed to quantify soluble TNFR1 in biological samples.

Key considerations for use as an ELISA standard:

• Purity and Formulation: The recombinant protein should be of high purity and formulated appropriately (often with carrier proteins like BSA for stability), as recommended for ELISA standards.
• Validation: It is essential to confirm that the recombinant TNFR1 behaves similarly to the native protein in your assay system. Commercial ELISA kits typically use recombinant TNFR1 as their calibration standard, indicating its suitability for quantification.
• Calibration Curve: Prepare a serial dilution of the recombinant TNFR1 to generate a standard curve, which allows for accurate quantification of TNFR1 in your samples.
• Activity and Epitope Recognition: Ensure that the recombinant protein contains the relevant epitopes recognized by the capture and detection antibodies in your ELISA. Most kits specify that both natural and recombinant TNFR1 are detected.

Limitations and best practices:

• Bioassay vs. ELISA: Recombinant proteins validated as ELISA standards may not be suitable for bioassays or functional studies unless specifically tested for those applications.
• Inactive Forms: ELISA may detect both biologically active and inactive forms of TNFR1, so quantification reflects total antigen rather than functional protein.
• Batch Consistency: Recombinant standards offer batch-to-batch consistency, which is critical for reproducible quantification.

Protocol summary:

1. Reconstitute the recombinant Mouse TNFR1 according to the manufacturer’s instructions.
2. Prepare serial dilutions in the recommended diluent to cover the expected concentration range.
3. Run the standard curve alongside your samples in the ELISA plate.
4. Use the standard curve to interpolate TNFR1 concentrations in unknown samples.

In summary, recombinant Mouse CD120a (TNFR1) is suitable as an ELISA standard for quantification, provided it is validated for your specific assay and prepared according to best practices for calibration.

Recombinant Mouse CD120a (TNFR1) has been validated for a range of scientific applications in published research, primarily for the detection, quantification, and functional analysis of TNFR1 in mouse models.

Validated Applications:

• Western Blot (WB): Used to detect TNFR1 protein expression in cell and tissue lysates.
• Immunohistochemistry (IHC): Applied to visualize TNFR1 localization in tissue sections, including mouse brain and cancer tissues.
• Immunofluorescence (IF-P): Enables detection of TNFR1 in fixed cells or tissue sections, such as mouse brain tissue.
• Enzyme-Linked Immunosorbent Assay (ELISA): Quantifies TNFR1 levels in biological samples.
• Immunoprecipitation (IP) and Co-Immunoprecipitation (CoIP): Used to isolate TNFR1 and study its protein-protein interactions.
• Flow Cytometry (FCM): Detects and quantifies TNFR1 on the surface of mouse cells.
• Functional Blocking/Neutralization Studies: Recombinant TNFR1 and its antibodies are used to block receptor signaling in vitro and in vivo, elucidating TNFR1’s role in immune regulation, apoptosis, and inflammation.
• In Vivo Studies: Functional grade antibodies targeting TNFR1 are used in mouse models to study autoimmune and inflammatory diseases, often by blocking receptor activity to assess physiological outcomes.

Research Contexts:

• Signal Transduction: TNFR1 is central to TNF-α signaling, mediating NF-κB activation, apoptosis, and inflammatory responses.
• Disease Models: Recombinant TNFR1 and its antibodies are widely used in models of cancer, autoimmune disorders, and inflammatory diseases to investigate therapeutic interventions and mechanistic pathways.
• Protein Interaction Studies: TNFR1’s interactions with adaptor proteins (e.g., TRADD, TRAF2, BAG4/SODD) are studied using immunoprecipitation and co-immunoprecipitation.

Summary Table of Validated Applications

These applications are supported by published research and validated protocols, making recombinant mouse CD120a (TNFR1) a versatile tool for immunological, cell biology, and disease model studies.

To reconstitute and prepare Recombinant Mouse CD120a (TNFR1) protein for cell culture experiments, follow these best practices based on standard protocols for recombinant proteins:

Reconstitution:

1. Centrifuge the vial before opening to ensure the lyophilized powder is at the bottom.
2. Reconstitute the protein in sterile distilled water to a final concentration of 0.1–0.5 mg/mL (some sources recommend up to 1.0 mg/mL). For example, add 100–500 µL of sterile water to 100 µg of protein.
3. Gently mix the solution by swirling or pipetting slowly. Avoid vortexing or vigorous pipetting to prevent protein denaturation.

Preparation for Cell Culture:

• If the protein will be used for long-term storage or repeated use, consider adding a carrier protein or stabilizer such as:
  • 0.1% BSA (bovine serum albumin)
  • 5% HSA (human serum albumin)
  • 10% FBS (fetal bovine serum)
  • 5% trehalose
• Aliquot the reconstituted protein into small working volumes to minimize freeze-thaw cycles.

Storage:

• Short-term storage: Store aliquots at 2–8°C for up to one month.
• Long-term storage: Store aliquots at -20°C to -80°C for up to 12 months.
• Avoid repeated freeze-thaw cycles, as this can reduce protein stability and activity.

Additional Notes:

• Always check the product-specific datasheet for any unique requirements, as some recombinant proteins may require specific buffers or additives.
• For cell culture applications, ensure all solutions are sterile and free of endotoxins.

By following these steps, you will help maintain the activity and stability of the Recombinant Mouse CD120a (TNFR1) protein for your experiments.