Recombinant Mouse Neuropoietin

Recombinant Mouse Neuropoietin is used in research applications primarily for its roles in neural cell survival, proliferation, and signaling, making it valuable for studies in neurobiology, neurodevelopment, and neuroprotection.

Key scientific reasons to use recombinant mouse neuropoietin include:

• Promotion of Embryonic Motor Neuron Survival: Neuropoietin supports the survival of embryonic motor neurons, which is critical for developmental neurobiology studies and for modeling neurodegenerative diseases affecting motor neurons.
• Stimulation of Neural Precursor Cell Proliferation: In the presence of growth factors such as EGF (epidermal growth factor) and FGF-2 (fibroblast growth factor 2), neuropoietin can increase the proliferation of neural precursor cells. This property is useful for research on neural stem cell biology, neurogenesis, and regenerative medicine.
• Unique Receptor Signaling: Neuropoietin signals through a receptor complex composed of CNTF receptor-α, gp130, and LIF receptor, activating downstream pathways such as STAT3. This makes it a tool for dissecting cytokine signaling mechanisms in neural cells.
• Modeling Species-Specific Functions: The human neuropoietin gene is suggested to have evolved toward a pseudogene, so mouse neuropoietin provides a unique opportunity to study functions that may not be present in humans, aiding in comparative neurobiology and evolutionary studies.

Additional considerations:

• Bioactivity Validation: Recombinant mouse neuropoietin is validated for bioactivity in cell-based assays, ensuring its suitability for in vitro and in vivo research applications.
• Research Use Only: It is intended strictly for research purposes and not for therapeutic or diagnostic use in humans.

In summary, use recombinant mouse neuropoietin in your research if you are investigating neural cell survival, neurodevelopment, cytokine signaling, or neural precursor proliferation, especially in mouse models where this cytokine is biologically active.

You can use recombinant Mouse Neuropoietin as a standard for quantification or calibration in ELISA assays, provided that the protein is sufficiently pure, its concentration is accurately determined, and it is compatible with the antibodies and detection system used in your assay.

Essential context and supporting details:

• Recombinant proteins are commonly used as standards in ELISA assays when purified native protein is unavailable. The key requirements are that the recombinant protein is well-characterized, pure, and its concentration is reliably measured (e.g., by absorbance at 280 nm, BCA assay, or HPLC).
• The standard curve in ELISA is generated by serial dilutions of the standard protein, spanning the expected concentration range of your samples. This allows for accurate quantification of unknowns by interpolation.
• Assay compatibility: The recombinant Neuropoietin must be recognized by the capture and detection antibodies in your ELISA. If your ELISA kit or protocol is designed for mouse Neuropoietin, recombinant protein is suitable as a calibrator, assuming it retains the relevant epitopes.
• Calibration and accuracy: Some commercial ELISA kits are calibrated against recombinant standards and validated for quantification of both recombinant and endogenous proteins. However, results may vary between kits and manufacturers, so it is important to validate recovery and parallelism in your specific assay system.
• Best practices: Prepare fresh dilutions of the recombinant standard in appropriate buffer, avoid protein-adsorbing plastics, and run the standard curve in parallel with your samples for each assay.

Additional relevant information:

• If your recombinant Neuropoietin is not supplied with a certificate of analysis or purity data, you should verify its concentration and purity before use.
• Always confirm that the recombinant protein behaves similarly to endogenous Neuropoietin in your assay, especially regarding antibody recognition and matrix effects.
• If available, refer to international standards for cytokines or growth factors for calibration, but recombinant protein is acceptable when such standards are not available.

In summary, recombinant Mouse Neuropoietin is suitable as an ELISA standard if it meets purity, quantification, and antibody recognition criteria, and you validate its performance in your assay system.

Recombinant Mouse Neuropoietin has been validated in published research primarily for applications related to bioactivity assays, neural cell survival and differentiation, and metabolic regulation in adipose tissue.

Key validated applications include:

• Promotion of embryonic motor neuron survival: Recombinant Mouse Neuropoietin has been shown to support the survival of embryonic motor neurons in vitro.
• Stimulation of neural precursor cell proliferation: In the presence of EGF and FGF-2, Neuropoietin increases the proliferation of neural precursor cells.
• Induction of astrocyte differentiation: Neuropoietin can induce neuroepithelial cells to differentiate into astrocytes, and acts synergistically with BMP2 to promote astrocyte differentiation.
• Inhibition of adipogenesis and metabolic effects: Neuropoietin blocks adipocyte differentiation and substantially attenuates insulin-stimulated glucose uptake in cultured adipocytes and murine adipose tissue, indicating a role in metabolic regulation.
• Bioactivity validation: The protein’s biological activity has been confirmed in cell proliferation assays using factor-dependent human erythroleukemic cell lines (e.g., TF-1 cells).
• Use as an ELISA standard: Recombinant Mouse Neuropoietin is also validated for use as a standard in ELISA assays for quantifying protein levels in research samples.

Supporting details:

• Neuropoietin acts through a receptor complex involving CNTF receptor-α, gp130, and LIF receptor, activating downstream STAT3 signaling.
• The protein is not expressed in humans, but is functionally relevant in mouse models for studying neurodevelopment and metabolic pathways.
• Validation for use in direct ELISAs and Western blots has been reported for antibodies raised against recombinant mouse Neuropoietin.

No published research was found validating its use in clinical or diagnostic applications; all uses are for basic and preclinical research. If you require details on a specific application (e.g., in vivo neuroprotection, metabolic disease models), please specify.

Reconstitution Procedures

Recombinant mouse neuropoietin (NP) is typically supplied as a lyophilized powder and requires proper reconstitution before use in cell culture experiments. The reconstitution concentration varies depending on the specific preparation and intended application.

Standard Reconstitution Protocol

Reconstitute the lyophilized protein in sterile phosphate-buffered saline (PBS) at a concentration of 0.2 mg/mL or 100 μg/mL, depending on your experimental requirements. Before opening the vial, briefly centrifuge it to bring the contents to the bottom. Gently reconstitute the powder without vortexing to minimize protein denaturation and foam formation. Some formulations may be reconstituted in sterile, distilled water at concentrations ranging from 0.1–1.0 mg/mL, so consult your specific product documentation for optimal parameters.

Formulation Considerations

The protein is typically lyophilized from a 0.2 μm filtered solution in PBS, often with trehalose as a cryoprotectant. Some preparations may contain alternative buffers such as 20 mM Tris-HCl (pH 8.0) with DTT and NaCl. The mature mouse neuropoietin protein has a molecular mass of approximately 20 kDa and typically contains approximately 80% methionyl form and 20% non-methionyl form.

Storage and Stability

Long-term Storage

Store the lyophilized protein desiccated at -20°C to -70°C in a manual defrost freezer for six to twelve months. Upon receipt, store the product immediately at the recommended temperature, as it is typically shipped at ambient temperature.

Post-Reconstitution Storage

After reconstitution under sterile conditions, the protein stability depends on temperature:

• At 2–8°C: stable for one month
• At -20 to -70°C: stable for three months

To prevent protein degradation, avoid repeated freeze-thaw cycles by preparing appropriate working aliquots. Store aliquots in sterile conditions to maintain sterility and prevent contamination.

Quality Parameters

The recombinant mouse neuropoietin preparation typically meets the following specifications:

• Purity: >97% by SDS-PAGE under reducing conditions, visualized by silver stain
• Endotoxin Level: <1.0 EU per 1 μg of protein, as determined by the LAL (Limulus Amebocyte Lysate) method
• Biological Activity: Determined in cell proliferation assays; the ED₅₀ for biological effects is typically 50–200 ng/mL

Application Considerations

Neuropoietin signals through a receptor complex comprised of CNTF receptor-α, gp130, and LIF receptor, with signaling involving STAT3. The protein promotes survival of embryonic motor neurons and can increase proliferation of neural precursor cells in the presence of EGF and FGF-2. For your specific cell culture experiments, determine optimal working concentrations empirically, as recommended dilutions vary by application and cell type.