Recombinant Human Leptin R

Using Recombinant Human Leptin R (Leptin Receptor) in research applications is essential for studying the molecular mechanisms of leptin signaling, its physiological effects, and its therapeutic potential in metabolic, endocrine, and neurobiological contexts.

Key reasons to use Recombinant Human Leptin R in research:

• Mechanistic Studies: The recombinant receptor enables precise investigation of leptin–receptor interactions, allowing researchers to dissect downstream signaling pathways involved in energy homeostasis, appetite regulation, and metabolic control.
• Bioassays and Binding Studies: Recombinant Leptin R is widely used in bioassays to quantify leptin activity, screen for agonists/antagonists, and characterize binding kinetics using techniques such as surface plasmon resonance.
• Therapeutic Research: Understanding how leptin binds and activates its receptor is critical for developing therapeutic strategies for obesity, lipodystrophy, and metabolic syndrome, as leptin signaling is central to these conditions.
• Modeling Human Disease: Recombinant Leptin R allows for the creation of in vitro models that mimic human leptin signaling, facilitating the study of mutations, receptor isoforms, and leptin resistance mechanisms relevant to human disease.
• Drug Discovery: The receptor is a valuable tool for screening small molecules or biologics that modulate leptin signaling, which could lead to novel treatments for metabolic and neuroendocrine disorders.

Supporting context:

• Leptin, via its receptor, regulates food intake, energy expenditure, neuroendocrine function, fertility, and immune responses.
• Recombinant leptin and its receptor have been used to demonstrate improvements in insulin resistance, lipid profiles, and body composition in both animal models and clinical studies, highlighting the translational value of receptor-based assays.
• The receptor is also used to study central nervous system effects of leptin, including cognition and mental health, by enabling targeted mechanistic experiments.

In summary, Recombinant Human Leptin R is a critical reagent for elucidating leptin biology, modeling disease, and advancing therapeutic research in metabolism and endocrinology.

Recombinant Human Leptin R (Leptin Receptor) can be used as a standard for quantification or calibration in ELISA assays, provided it is well-characterized, matches the isoform detected by your assay, and is compatible with your assay’s antibodies and detection system. This is a common practice in quantitative ELISA development and is supported by multiple technical resources.

Key considerations and supporting details:

• Assay Compatibility: Commercial ELISA kits for human Leptin R (Leptin Receptor) routinely use recombinant human Leptin R as the standard for generating calibration curves. These standards are typically supplied as lyophilized recombinant protein, reconstituted and serially diluted to create a standard curve for quantification.

• Isoform Specificity: The Leptin receptor exists in multiple isoforms (e.g., long and short forms). Ensure that the recombinant standard you use matches the isoform recognized by your assay’s capture and detection antibodies. Most commercial kits specify which isoform their antibodies detect.

• Validation: The recombinant standard should be validated for use in your specific assay format. Commercial kits demonstrate that their recombinant Leptin R standard yields dose-response curves parallel to those obtained with natural Leptin R, indicating suitability for quantification. If using a recombinant standard from a different source, confirm parallelism and recovery in your matrix.

• Traceability: Some kits calibrate their standards against international reference materials (e.g., NIBSC/WHO standards for Leptin, though not always available for Leptin R). If traceability to a reference standard is required for your application, check if your recombinant Leptin R is traceable or comparable.

• Documentation: For publication or regulatory purposes, document the source, purity, and characterization of your recombinant standard, and validate its performance in your assay.

Protocol best practices:

• Prepare a serial dilution of the recombinant Leptin R in the same buffer/matrix as your samples to generate a standard curve.
• Run the standard curve in parallel with your samples on each plate.
• Validate linearity, recovery, and parallelism between the recombinant standard and endogenous Leptin R in your sample matrix.

Limitations:

• If your recombinant Leptin R differs significantly in glycosylation or structure from the endogenous protein, this may affect antibody recognition and quantification accuracy. Always verify assay performance with your chosen standard.

In summary, using recombinant human Leptin R as a standard is standard practice in quantitative ELISA assays for Leptin R, provided you confirm compatibility and validate its performance in your specific assay system.

Recombinant Human Leptin Receptor (Leptin R) has been validated for several key applications in published research, primarily in biochemical and cell-based assays.

Validated Applications:

• Bioassays: Used to assess the biological activity of leptin and its receptor interactions, including cell proliferation assays in leptin-responsive cell lines.
• Binding Assays: Employed to study the binding affinity and kinetics between leptin and its receptor, often using recombinant proteins.
• Surface Plasmon Resonance (SPR): Utilized for real-time analysis of leptin-leptin receptor interactions, providing quantitative binding data.
• ELISA (Standard): Used as a standard or capture reagent in enzyme-linked immunosorbent assays to quantify leptin or leptin receptor levels.
• In Vivo Studies: Applied in animal models to investigate leptin receptor function and leptin signaling pathways.
• Neutralization Assays: Recombinant Leptin R has been used to neutralize the activity of recombinant human leptin in leptin-dependent cell systems, confirming its functional activity.

Supporting Details:

• Cellular Bioassays: Recombinant Human Leptin R is frequently used in cell-based assays to validate leptin-induced signaling, proliferation, migration, and cytokine production in various human and mouse cell types.
• Protein-Protein Interaction Studies: SPR and other binding assays with recombinant Leptin R help characterize the molecular interactions and specificity of leptin binding.
• ELISA Standards: Recombinant Leptin R serves as a standard for quantification in immunoassays, supporting studies of leptin receptor expression and function.
• Functional Neutralization: The ability of recombinant Leptin R to neutralize leptin activity is a critical validation for its use in mechanistic studies of leptin signaling.

Additional Notes:

• Recombinant Human Leptin R is typically expressed as a soluble extracellular domain, often fused to Fc or His tags for purification and detection purposes.
• It is used in both basic research and preclinical studies to dissect leptin receptor biology, including metabolic, endocrine, and immunological functions.

Summary Table:

These applications are well-supported in published research and are foundational for studies of leptin signaling, receptor pharmacology, and metabolic regulation.

To reconstitute and prepare Recombinant Human Leptin R (Leptin Receptor) protein for cell culture experiments, follow these best-practice steps:

• Centrifuge the vial briefly before opening to ensure all lyophilized protein is at the bottom.
• Reconstitute the protein in sterile buffer. The most common recommendations are:
  • Use sterile distilled water or sterile PBS (phosphate-buffered saline).
  • Target a final concentration of 0.1–0.5 mg/mL. Some protocols recommend up to 200 μg/mL in PBS.
• Avoid vigorous mixing: Do not vortex or pipette vigorously, as this can denature the protein. Instead, gently swirl or invert the vial to dissolve.
• Stabilize the protein for cell culture use:
  • For long-term storage or to prevent adsorption/loss, add a carrier protein such as 0.1% BSA (bovine serum albumin), 5% HSA (human serum albumin), 10% FBS (fetal bovine serum), or 5% trehalose.
• Aliquot the reconstituted solution to avoid repeated freeze-thaw cycles, which can degrade the protein.
• Storage:
  • Short-term: Store at 2–8°C for up to 1 week.
  • Long-term: Store aliquots at –20°C to –80°C.
• Working solution: Before adding to cell culture, dilute the protein in your cell culture medium or appropriate buffer to the desired working concentration, ensuring compatibility with your assay and cell type.

Example protocol for reconstitution:

1. Briefly centrifuge the vial.
2. Add sterile PBS (or distilled water) to achieve 0.1–0.5 mg/mL.
3. Gently mix by swirling or inverting; let sit at room temperature for 10–30 minutes to fully dissolve.
4. If needed, add 0.1% BSA for stability.
5. Aliquot and store as described above.

Additional notes:

• Always consult the specific product datasheet for any unique requirements, as buffer composition and concentration may vary by manufacturer or protein construct.
• For cell-based assays, ensure the final buffer is compatible with your cells (e.g., avoid high salt or non-physiological pH).

These steps will help maintain the bioactivity and stability of recombinant human Leptin R protein for reliable cell culture experiments.