Recombinant Mouse Leptin

Recombinant Mouse Leptin is widely used in research to study the physiological and pathological roles of leptin in metabolism, energy homeostasis, immune function, and disease models, particularly in mice. Its use is essential for controlled, reproducible experiments investigating leptin signaling and deficiency.

Key reasons to use recombinant mouse leptin in research applications:

• Modeling Leptin Deficiency and Replacement: Recombinant mouse leptin is critical for studying the effects of leptin replacement in leptin-deficient (ob/ob) mice, which are genetically obese due to mutations in the leptin gene. Administration of recombinant leptin to these mice reverses obesity and corrects associated metabolic and neuroendocrine abnormalities, providing a direct tool to dissect leptin’s physiological functions.

• Metabolic and Endocrine Research: Leptin regulates appetite, energy expenditure, glucose metabolism, and lipid homeostasis. Recombinant leptin allows researchers to manipulate leptin levels in vivo or in vitro, enabling studies on metabolic diseases such as obesity, diabetes, and lipodystrophy.

• Immunology and Inflammation: Leptin modulates immune cell function, including T-cell differentiation and macrophage activation. Recombinant leptin is used in bioassays and cell culture to investigate its immunomodulatory effects and its role in autoimmune and inflammatory diseases.

• Neurobiology: Leptin influences neuroendocrine pathways, synaptic plasticity, and hypothalamic regulation of energy balance. Recombinant leptin administration in mice is used to study these central nervous system effects, including changes in neuronal connectivity and neuropeptide expression.

• Therapeutic and Translational Studies: Recombinant leptin is used to test potential therapeutic interventions for rare conditions such as congenital leptin deficiency and lipodystrophy, and to explore its effects in models of metabolic syndrome and insulin resistance.

• Experimental Control: Using recombinant protein ensures precise dosing, batch-to-batch consistency, and the ability to perform dose-response studies, which are critical for reproducible and interpretable results in both in vivo and in vitro experiments.

Common applications include:

• In vivo administration to mice for metabolic, immunological, or neurobiological studies.
• In vitro assays with primary cells or cell lines to assess leptin signaling and downstream effects.
• Bioassays to evaluate leptin’s impact on immune cell differentiation, cytokine production, or metabolic activity.

In summary, recombinant mouse leptin is an indispensable tool for dissecting leptin’s multifaceted roles in physiology and disease, enabling mechanistic studies and translational research in mouse models.

Yes, recombinant mouse Leptin can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity and its concentration is accurately determined. This approach is widely accepted in research applications for the quantification of Leptin in biological samples.

Supporting details:

• ELISA kits for mouse Leptin commonly use recombinant mouse Leptin, typically expressed in E. coli, as the standard for generating calibration curves. These standards are validated to produce parallel dose-response curves when compared to natural mouse Leptin, indicating their suitability for quantification.
• The accuracy of quantification depends on the purity and precise concentration of the recombinant protein. It is essential to use a well-characterized recombinant Leptin preparation, ideally calibrated against an international reference standard when available.
• Protocols: Prepare a dilution series of the recombinant Leptin standard in the same buffer as your samples (often a calibrator diluent provided with the kit or a suitable buffer such as PBS with carrier protein), and use these dilutions to generate a standard curve. Sample concentrations are then interpolated from this curve.
• Limitations: The recombinant standard should closely match the native protein in terms of immunoreactivity. Most commercial recombinant mouse Leptin proteins are validated for this purpose, but it is advisable to confirm that your specific preparation is suitable for ELISA calibration, especially if using a custom or non-commercial source.

Best practices:

• Use recombinant mouse Leptin that is validated for ELISA applications.
• Confirm the protein’s concentration using an independent method (e.g., UV absorbance, BCA assay).
• Prepare standards freshly or aliquot and store appropriately to avoid degradation.
• Ensure the standard curve covers the expected range of sample concentrations.

Summary: Recombinant mouse Leptin is a scientifically accepted standard for ELISA quantification, provided it is properly validated and prepared. This enables accurate calibration and quantification of Leptin in mouse biological samples.

Recombinant Mouse Leptin has been validated for a wide range of applications in published research, primarily in in vivo studies, bioassays, and functional studies involving metabolic, immunological, and neurobiological processes.

Key validated applications include:

• In Vivo Studies:

  • Used to investigate body weight regulation, energy balance, and obesity models in mice, including leptin-deficient (ob/ob) and diet-induced obese models.
  • Studied for its effects on glucose homeostasis, insulin resistance, and glycemic regulation.
  • Applied in models of neurodegenerative diseases (e.g., ALS, Alzheimer’s) to assess neuroprotective effects and motor function improvement.
  • Used to explore roles in reproduction, hematopoiesis, and pubertal development.
  • Investigated for immune modulation, including effects on T-cell metabolism and differentiation, and in autoimmune disease models.

• Bioassays and In Vitro Functional Studies:

  • Employed to assess leptin receptor activity and downstream signaling in cell lines and primary cells.
  • Used to study gene expression changes in adipose tissue, particularly genes involved in inflammation and oxidative stress.
  • Applied in assays measuring chemokine expression in cultured macrophages and other immune cells.
  • Utilized in studies of angiogenesis, wound healing, and inflammatory responses.

• ELISA Standard and Detection:

  • Used as a standard in ELISA assays for quantifying leptin levels in biological samples.

• Other Applications:

  • Intranasal and intracerebroventricular delivery to study central nervous system effects and brain-fat axis signaling.
  • Functional studies in hematopoietic and lymphohematopoietic cell populations.

Summary Table of Validated Applications

Notable Published Examples:

• ALS mouse models: Intranasal recombinant mouse leptin improved motor performance and delayed disease progression.
• Obesity and diabetes models: Used to study insulin and leptin resistance, and to restore metabolic balance in leptin-deficient mice.
• Immunology: Demonstrated direct effects on T-cell metabolism and differentiation, and modulation of immune responses in autoimmunity and infection.

These applications are well-supported by peer-reviewed studies and product validation data, confirming the broad utility of recombinant mouse leptin in both basic and translational research contexts.

To reconstitute and prepare Recombinant Mouse Leptin protein for cell culture experiments, follow these general guidelines based on manufacturer recommendations and best practices:

Reconstitution

1. Centrifuge the vial briefly before opening to ensure all lyophilized powder is at the bottom.

2. Reconstitute the lyophilized protein in sterile, endotoxin-free water or an appropriate buffer. Common recommendations include:

   • Sterile distilled water (e.g., 1.0–5.0 mg/mL, as per Thermo Fisher).
   • Sterile 20 mM Tris-HCl, pH 8.0 (as per R&D Systems).
   • Sterile 18 MΩ-cm H₂O (as per Prospec Bio).
   • For some products, reconstitution in 20 mM HCl may be recommended (e.g., Abcam ab222365).

   Note: Always check the specific product datasheet for the recommended buffer and concentration.

3. Gently mix the solution until fully dissolved. Avoid vigorous shaking to prevent denaturation.

Preparation for Cell Culture

1. Aliquot the reconstituted protein into small working volumes to minimize freeze-thaw cycles.
2. Store aliquots at –20°C to –80°C for long-term storage. For short-term use, store at 2–8°C for up to 1 month.
3. Avoid repeated freeze-thaw cycles to maintain protein activity.
4. Dilute the protein in cell culture medium or an appropriate buffer before adding to cells. For optimal stability, dilutions can be made in aqueous buffers supplemented with carrier protein (e.g., 0.1–1.0% BSA or HSA), especially if the protein will be stored after dilution.

Additional Tips

• Sterile conditions are essential for cell culture applications.
• Carrier proteins (e.g., BSA or HSA) can help stabilize the leptin in solution, especially for prolonged storage or dilute solutions.
• Equilibrate the protein to room temperature before use to avoid precipitation.

Example Protocol

• Reconstitute 100 µg of lyophilized mouse leptin in 100 µL sterile water (1 mg/mL).
• Aliquot into 10 µL volumes and store at –80°C.
• For cell culture, thaw an aliquot, dilute in culture medium to the desired concentration (e.g., 10–100 ng/mL), and add to cells.

Always refer to the specific product datasheet for detailed instructions and recommended concentrations for your experimental setup.