Recombinant Mouse Prolactin

Recombinant Mouse Prolactin (mPRL) is a valuable tool for a wide range of research applications due to its well-characterized biological activity and versatility. Here are several key reasons why you should consider using recombinant mouse prolactin in your research:

1. Biological Relevance and Functional Activity

Recombinant mouse prolactin retains the functional properties of native prolactin, including its ability to:

• Stimulate mammary gland development and lactation.
• Promote cell proliferation in various cell types, such as Nb2 pre-T cell lymphoma cells and adipocytes.
• Modulate immune responses and lymphoid tissue development.
• Influence adipocyte differentiation and gene expression (e.g., upregulation of PPARγ and downregulation of C/EBPβ and FAS).

These activities make it suitable for studying physiological and pathological processes related to prolactin signaling in mice and other rodent models.

2. Consistency and Purity

Recombinant prolactin is produced under controlled conditions, ensuring high batch-to-batch consistency and purity (often >95–99% as determined by SDS-PAGE and Western blot). This reduces experimental variability and enhances reproducibility in your studies.

3. Flexibility in Experimental Design

Recombinant mouse prolactin can be used in a variety of experimental settings, including:

• In vitro assays: Cell proliferation, differentiation, signaling, and receptor binding studies.
• In vivo studies: Animal models of lactation, immune modulation, and metabolic regulation.
• Mechanistic studies: Investigation of prolactin receptor (PRLR) signaling pathways, such as JAK-STAT activation.

4. Cost-Effectiveness and Scalability

Recombinant production in systems like E. coli or insect cells allows for scalable and cost-effective manufacturing, making it accessible for both small-scale laboratory experiments and larger studies.

5. Support for Translational Research

Mouse prolactin shares significant homology with human prolactin, allowing findings from mouse models to inform understanding of human physiology and disease. This is particularly useful for studies on lactation, immune regulation, and metabolic disorders.

6. Well-Validated Applications

Recombinant mouse prolactin has been successfully used in published studies to:

• Investigate the role of prolactin in adipocyte differentiation and metabolism.
• Study immune cell proliferation and cytokine-like activities.
• Explore neuroendocrine functions and hippocampal precursor cell proliferation.

7. Ease of Use and Storage

Most recombinant mouse prolactin products are supplied as lyophilized powders or concentrated solutions, which are stable and easy to reconstitute and store, minimizing handling and degradation issues.

In summary, recombinant mouse prolactin is a reliable, biologically active, and versatile reagent that supports a broad spectrum of research applications in endocrinology, immunology, metabolism, and developmental biology. Its use enables precise, reproducible, and mechanistic investigations into prolactin’s diverse roles in health and disease.

Yes, recombinant mouse prolactin can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated and matched to the assay system. Recombinant prolactin is commonly used as a calibrator in mouse prolactin ELISA kits, and most commercial kits are designed to recognize both native and recombinant forms of the protein.

Key considerations for use:

• Assay Compatibility: Ensure your ELISA kit is validated to detect recombinant mouse prolactin. Most sandwich ELISA kits for mouse prolactin specify that their antibodies recognize both native and recombinant forms, and their standard curves are often generated using recombinant protein.
• Purity and Activity: The recombinant prolactin should be of high purity (typically >95% by SDS-PAGE/HPLC) and retain biological activity, as confirmed by the manufacturer or your own QC.
• Formulation: Recombinant standards may be supplied lyophilized or in solution, sometimes with carrier proteins (e.g., BSA) to stabilize them. Follow recommended reconstitution protocols to ensure consistency.
• Calibration Curve: Prepare a serial dilution of the recombinant standard in the same buffer or diluent as your samples to generate a calibration curve. This allows accurate quantification of prolactin in unknown samples.
• Validation: It is best practice to validate the recombinant standard in your specific ELISA system, confirming parallelism between the standard curve and endogenous sample dilution curves. This ensures the recombinant protein behaves similarly to native prolactin in your assay conditions.

Limitations:

• Minor differences in glycosylation or folding between recombinant and native prolactin may affect antibody recognition in rare cases, but most commercial ELISA kits are designed to minimize such issues.
• Always check the kit documentation for any specific recommendations or restrictions regarding standard protein source.

Summary Table: Use of Recombinant Mouse Prolactin as ELISA Standard

In conclusion, recombinant mouse prolactin is widely accepted and scientifically appropriate as a standard for ELISA quantification and calibration, provided you follow best practices for validation and preparation.

Recombinant Mouse Prolactin has been validated for several key applications in published research, primarily in cell-based assays, in vivo studies, and molecular analyses.

Validated Applications:

• Cell Differentiation and Signaling Assays: Recombinant mouse prolactin has been used to study its effects on the differentiation of 3T3-L1 adipocytes, including analysis of mRNA expression for genes such as C/EBPβ, FAS, and PPARγ, confirming its biological activity in regulating adipocyte differentiation and gene expression.
• Cytotoxicity Assays: Its impact on cell viability has been assessed using standard cytotoxicity assays (e.g., CCK-8) in 3T3-L1 cells, demonstrating no cytotoxic effects at tested concentrations.
• Bioassays (Functional Activity): The protein has been validated in bioassays to stimulate precursor cells in the adult mouse hippocampus, indicating its role in neurogenesis and cell proliferation.
• In Vivo Studies: Recombinant mouse prolactin has been used in animal models to investigate physiological roles such as lactation, metabolic adaptations during pregnancy, and pain sensitization, including studies on prolactin receptor signaling and nociceptor sensitization.
• Western Blot and SDS-PAGE: Its identity and purity have been confirmed by western blot and SDS-PAGE, validating its use in protein detection and quantification protocols.
• Receptor Binding Assays: The protein has been used to study prolactin receptor interactions and downstream signaling pathways in various cell types.
• ELISA Standard: Recombinant mouse prolactin is commonly used as a standard in ELISA assays for quantifying prolactin levels in biological samples.

Additional Context:

• Immune Function Studies: Research has explored prolactin’s role in immune regulation, including its effects on immune cell function and inflammatory responses.
• Neuroendocrine Research: Prolactin’s influence on neuroendocrine pathways, especially in the context of lactation and reproductive biology, has been validated in both cell culture and animal models.
• Protein Characterization: Analytical techniques such as HPLC and SDS-PAGE have been used to confirm the purity and structural integrity of recombinant mouse prolactin.

Summary Table of Validated Applications

These applications are supported by peer-reviewed studies and product validation data, confirming the utility of recombinant mouse prolactin in diverse research areas including endocrinology, metabolism, neurobiology, and immunology.

To reconstitute and prepare Recombinant Mouse Prolactin protein for cell culture experiments, dissolve the lyophilized protein in sterile buffer to a concentration of at least 100 μg/mL, using either sterile water or PBS, and include a carrier protein such as BSA if required for stability.

Step-by-step protocol:

• Centrifuge the vial briefly before opening to ensure all lyophilized material is at the bottom.
• Reconstitution:
  • Add sterile ultrapure water (18 MΩ-cm) or sterile PBS to achieve a final concentration of at least 100 μg/mL.
  • If the protein is prone to adsorption or instability, add 0.1% BSA (bovine serum albumin) as a carrier to the buffer.
  • Gently mix by pipetting or slow inversion; avoid vigorous shaking or vortexing to prevent protein denaturation or foaming.
  • Allow the protein to fully dissolve at room temperature for 15–30 minutes with gentle agitation.
• Aliquot the reconstituted solution to avoid repeated freeze-thaw cycles, which can degrade the protein.
• Storage: Use immediately for cell culture or store aliquots at 4°C for up to a week, or at –20°C/–80°C for long-term storage.
• Dilution for cell culture: Further dilute the stock solution to the desired working concentration using sterile cell culture medium or buffer immediately before use.

Additional notes:

• Always use sterile technique to prevent contamination.
• If using for sensitive cell types or serum-free culture, ensure all reagents (including BSA) are cell culture grade and endotoxin-free.
• The optimal working concentration for cell culture will depend on your specific assay and cell type; typical ranges are 1–100 ng/mL, but titration may be necessary.

Summary of key points:

• Reconstitute at ≥100 μg/mL in sterile water or PBS, with 0.1% BSA if needed.
• Mix gently and allow to dissolve completely.
• Aliquot and store appropriately to maintain protein integrity.
• Dilute to working concentration in cell culture medium just before use.

This protocol ensures the recombinant prolactin is properly solubilized and stable for use in cell culture experiments.