Recombinant Human Prolactin Receptor

Using Recombinant Human Prolactin Receptor (PRLR) in research applications enables precise investigation of prolactin signaling, receptor-ligand interactions, and downstream biological effects relevant to lactation, reproduction, immunology, and cancer biology.

Key scientific advantages and applications:

• Mechanistic Studies: Recombinant PRLR allows controlled analysis of prolactin binding, receptor activation, and signal transduction pathways in vitro, facilitating dissection of molecular mechanisms underlying prolactin’s effects on target cells.
• Functional Assays: It enables quantitative assays of ligand (prolactin or analogs) activity, including measurement of receptor-mediated gene expression (e.g., activation of milk protein promoters), cell proliferation, or differentiation.
• Drug Discovery and Screening: Recombinant PRLR is essential for screening and characterizing prolactin receptor modulators (agonists, antagonists, or neutral ligands), which are being explored for therapeutic applications in lactation disorders, reproductive health, and oncology.
• Cancer Research: Altered PRLR expression and signaling are implicated in breast and prostate cancer progression. Recombinant PRLR can be used to model these changes, study receptor isoform-specific effects, and test targeted inhibitors.
• Immunological Studies: PRLR is involved in immune regulation; recombinant receptor enables investigation of prolactin’s immunomodulatory roles and receptor-mediated signaling in immune cells.

Technical benefits:

• Consistency and Purity: Recombinant PRLR provides a standardized, high-purity reagent, minimizing variability and background noise compared to native or tissue-derived receptors.
• Customization: Recombinant technology allows for engineering of receptor isoforms, mutations, or tagged versions (e.g., His-tag), supporting structure-function studies and facilitating detection or purification.

Summary of best practices:

• Use recombinant PRLR in cell-based assays, binding studies, or biochemical analyses to ensure reproducibility and specificity.
• Select appropriate receptor isoforms or mutants to address specific research questions (e.g., short vs. long forms in cancer studies).
• Validate receptor functionality and ligand specificity in your assay system.

In conclusion, recombinant human prolactin receptor is a critical tool for elucidating prolactin biology, developing therapeutics, and advancing translational research in endocrinology, reproduction, immunology, and oncology.

You can use recombinant human prolactin receptor (PRLR) protein as a standard for quantification or calibration in ELISA assays, provided the assay is specifically designed to detect PRLR and validated for use with recombinant protein standards.

Key considerations:

• Assay Compatibility: The ELISA must be validated to recognize both recombinant and native PRLR. Many commercial PRLR ELISA kits use recombinant PRLR as their standard, enabling quantification of PRLR in biological samples. Always confirm that your assay’s antibodies recognize the recombinant form you intend to use, as some kits are designed only for native PRLR.

• Standard Preparation: Recombinant PRLR standards should be prepared according to the manufacturer’s instructions, typically by reconstituting the lyophilized protein and diluting to the desired concentration range for your standard curve. Storage conditions (e.g., -20°C or -80°C) and handling protocols must be followed to maintain protein integrity.

• Validation: It is essential to validate the linearity, recovery, and parallelism of your standard curve using the recombinant PRLR. This ensures that quantification in your samples is accurate and comparable to the standard.

• Biological Activity: Note that some recombinant PRLR preparations are not biologically active, which is generally acceptable for quantification purposes in ELISA, as the assay detects antigenic epitopes rather than functional activity.

Limitations and Best Practices:

• Isoform and Epitope Specificity: Recombinant PRLR may represent a specific isoform or fragment. Ensure that the standard matches the isoform or domain detected by your ELISA antibodies.
• Matrix Effects: When quantifying PRLR in complex biological matrices (e.g., serum, plasma), confirm that the recombinant standard behaves similarly to endogenous PRLR in your assay system (parallelism testing).

Summary Table: Recombinant PRLR as ELISA Standard

If your ELISA is designed for prolactin (PRL) quantification, you must use recombinant or native PRL as the standard, not PRLR. PRLR standards are only appropriate for assays measuring the receptor itself.

In summary: Recombinant human PRLR can be used as a standard for ELISA quantification of PRLR, provided the assay is validated for this purpose and the standard is properly prepared and matched to the assay’s detection antibodies.

Recombinant Human Prolactin Receptor (PRLR) has been validated for several key applications in published research, primarily in the fields of cancer biology, cell signaling, and receptor-ligand interaction studies.

Validated Applications:

• Cell-based Bioassays: Recombinant PRLR is widely used in bioassays to study prolactin signaling, including activation of downstream pathways such as STAT5 phosphorylation in breast and prostate cancer cell lines. These assays help elucidate the functional consequences of PRLR activation or inhibition.

• Cancer Research: PRLR is a target in hormone-dependent cancers (breast, prostate, ovarian, endometrial). Recombinant PRLR and its antagonists (e.g., Δ1–9-G129R-hPRL) have been used to validate therapeutic strategies that block PRLR signaling, demonstrating effects such as inhibition of tumor growth, suppression of proliferation, and reduction of chemoresistance in various cancer models.

• Ligand Binding and Receptor Characterization: Recombinant PRLR is used to characterize binding affinities and specificities of prolactin and its antagonists, as well as to screen for neutralizing antibodies or small molecule inhibitors targeting the receptor.

• Protein-Protein Interaction Studies: Recombinant PRLR enables investigation of interactions with prolactin and other ligands, facilitating studies on receptor activation, dimerization, and downstream signaling events.

• Cell Differentiation and Signaling: Recombinant PRLR is used in vitro to study its role in cell differentiation, especially in mammary epithelial cells and immune cells, and to dissect the molecular mechanisms of prolactin-induced signaling.

Supporting Details:

• Cancer Therapeutics: Modified prolactin antagonists (e.g., G129R-hPRL, Δ1–9-G129R-hPRL) have been validated using recombinant PRLR to block receptor signaling in cancer cell lines and animal models, showing reduced tumor growth and enhanced therapeutic efficacy when combined with other agents.

• Antibody Development: Recombinant PRLR is essential for validating the specificity and efficacy of prolactin-neutralizing monoclonal antibodies, which are being developed for conditions such as female-selective pain and endometriosis.

• Receptor Overexpression Systems: Recombinant PRLR expressed in HEK293 or E. coli systems is used to generate cell lines or supernatants for functional assays, ligand screening, and mechanistic studies.

Summary Table:

Note: While recombinant PRLR is primarily used in vitro, its validation in published research spans diverse experimental systems, including cell lines, recombinant protein assays, and animal models. The receptor is a central tool for dissecting prolactin biology and developing targeted therapies for cancer and other prolactin-related disorders.

To reconstitute and prepare Recombinant Human Prolactin Receptor (PRLR) protein for cell culture experiments, follow these steps for optimal solubility and biological activity:

1. Centrifuge the vial briefly before opening to ensure all lyophilized protein is at the bottom.
2. Reconstitution buffer: Use sterile deionized water or 10 mM PBS (pH 7.4), depending on your downstream application and the protein’s formulation. PBS is generally preferred for cell culture to maintain physiological pH and osmolarity.
3. Concentration: Reconstitute to a final concentration of 0.1–1.0 mg/mL. For most protocols, 0.1–0.4 mg/mL is recommended.
4. Mixing: Gently pipette up and down to dissolve the protein. Do not vortex, as vigorous mixing can denature or aggregate the protein.
5. Carrier protein (optional): If long-term storage or repeated freeze-thaw cycles are expected, consider adding a carrier protein such as 0.1% BSA to stabilize the recombinant PRLR. This is optional and should be avoided if BSA may interfere with your assay.
6. Aliquoting and storage: After reconstitution, aliquot the solution to avoid repeated freeze-thaw cycles. Store aliquots at –80°C for long-term use, or at 2–8°C for short-term use (up to one month).
7. Sterility: Always use sterile technique and solutions to prevent contamination, especially for cell culture applications.

Protocol summary:

1. Briefly centrifuge the vial.2. Add sterile deionized water or PBS (pH 7.4) to achieve 0.1–1.0 mg/mL.3. Gently pipette to dissolve; do not vortex.4. Optional: Add 0.1% BSA for stability.5. Aliquot and store at –80°C (long-term) or 2–8°C (short-term).

Additional notes:

• If the protein is tagged (e.g., His-tag), ensure compatibility with your cell culture system and downstream assays.
• For functional assays, further dilute the reconstituted stock in cell culture medium immediately before use.
• Always verify the protein’s concentration using a protein assay after reconstitution if precise dosing is required.

This protocol ensures maximal recovery and stability of the recombinant PRLR protein for cell culture experiments.