Recombinant Human Procalcitonin

Recombinant human procalcitonin (hPCT) offers several compelling advantages for research and diagnostic applications, making it a valuable tool across multiple scientific domains.

Primary Research Applications

Biomarker Development and Validation

Recombinant hPCT serves as an essential reference standard for developing and validating diagnostic assays. It enables researchers to establish traceability to SI units through high-resolution mass spectrometry and amino acid analysis, ensuring accurate quantification in clinical and research settings. This standardization is critical for creating certified reference materials that support external quality assessment schemes and improve harmonization across different measurement platforms.

Immunoassay Development

The protein functions as an antigen for immunodetection applications, making it suitable for ELISA, Western blot, and other immunological techniques. Researchers can use recombinant hPCT to develop and optimize antibody-based detection methods, establish calibration curves, and validate assay performance across different platforms.

Diagnostic Platform Innovation

Recombinant hPCT enables the development of novel diagnostic platforms, including point-of-care testing systems. For example, fluorescence-based lateral flow assays have been successfully developed for simultaneous quantification of procalcitonin alongside other biomarkers. This allows researchers to create rapid, multiplexed diagnostic tools for clinical applications.

Technical Advantages

Expression and Purification

Recombinant hPCT can be efficiently expressed in bacterial systems such as E. coli T7 express strains, yielding tag-free protein with high purity (>95%). This bacterial expression system provides cost-effective production while maintaining biological activity and reducing contamination risks associated with endotoxins through careful purification protocols.

Protein Characteristics

The recombinant protein is typically produced as a single, non-glycosylated polypeptide chain containing 116 amino acids with a molecular mass of approximately 13 kDa. This consistent composition ensures reproducibility across experiments and batches, which is essential for standardization in research applications.

Clinical Research Support

Recombinant hPCT supports research aimed at improving antibiotic stewardship and sepsis diagnosis. By serving as a primary calibrator and reference material, it enables the development of more accurate and comparable immunoassays that can guide clinical decision-making. This contributes to research efforts focused on reducing unnecessary antibiotic exposure while maintaining patient safety and improving treatment outcomes.

Yes, recombinant human procalcitonin is commonly used as a standard for quantification or calibration in ELISA assays for procalcitonin measurement.

Supporting details:

• ELISA kits for human procalcitonin routinely use recombinant human procalcitonin as the standard to generate the calibration curve, enabling quantification of procalcitonin in biological samples such as plasma, serum, and cell culture supernatants.
• The recombinant standard is typically produced in E. coli and may represent the full-length or a specific fragment of procalcitonin (e.g., amino acids 1-126 or 26-141).
• The standard must be well-characterized, with known purity and concentration, to ensure traceability and accuracy in quantification. For high-precision applications, characterization by mass spectrometry and amino acid analysis is recommended.
• When preparing your standard curve, ensure the recombinant protein matches the epitope recognized by the antibodies in your ELISA kit (full-length or specific fragment), as this affects assay specificity and quantification.

Best practices:

• Reconstitute and dilute the recombinant standard according to the ELISA kit instructions to cover the assay’s dynamic range (e.g., 27.43–20,000 pg/mL).
• Validate recovery and linearity by spiking known amounts of recombinant procalcitonin into your sample matrix (serum, plasma, etc.).
• Confirm that the recombinant standard is compatible with your assay’s antibodies and detection method, as some kits specify the exact fragment or tag used.

Limitations:

• Results may vary between kits due to differences in antibody specificity, standard protein sequence, and calibration procedures. Standardization efforts are ongoing to harmonize procalcitonin measurements across platforms.
• For clinical or diagnostic applications, ensure your calibration procedure meets regulatory and traceability requirements.

In summary, recombinant human procalcitonin is suitable and widely accepted as a standard for ELISA quantification, provided it is appropriately characterized and matches your assay’s requirements.

Recombinant Human Procalcitonin has been validated in published research primarily for use as a calibrator and reference material in quantitative assays, as well as for the development and validation of diagnostic methods for detecting procalcitonin in human samples.

Key validated applications include:

• Primary calibrator in quantitative assays: Recombinant human procalcitonin is used as a standard to establish traceability in high-resolution mass spectrometry and amino acid analysis, supporting the absolute quantification of PCT in human serum. This enables assignment of target values to secondary certified reference materials, which are essential for external quality assessment and harmonization of commercial immunoassays.

• Positive control in immunoassays: It serves as a positive control for assay validation, ensuring accuracy and reliability in methods such as ELISA, chemiluminescent immunoassays, and other antibody-based detection platforms.

• Development and validation of rapid detection methods: Recombinant PCT has been used in the development of rapid detection techniques, including MALDI-TOF mass spectrometry and proximity extension assays (PEA), which are designed for sensitive and specific identification of bacterial infections and sepsis in clinical samples.

• Clinical research on biomarker utility: While recombinant PCT itself is not directly used in patient diagnosis, its validated use in assay development underpins clinical studies that utilize PCT as a biomarker for early detection of systemic bacterial infections, sepsis, and for guiding antibiotic stewardship in various clinical settings.

Additional relevant details:

• Traceability to SI units: Characterization of recombinant PCT by mass spectrometry and amino acid analysis ensures that assay results are traceable to international standards, improving comparability across laboratories and studies.
• External quality assessment: Its use in reference measurement procedures supports the monitoring and improvement of commercial assay accuracy, which is critical for reliable clinical decision-making.

In summary, recombinant human procalcitonin is validated for use as a calibrator, reference material, and positive control in the development, standardization, and validation of diagnostic assays for procalcitonin, which is a key biomarker in sepsis and bacterial infection research and clinical practice.

To reconstitute and prepare Recombinant Human Procalcitonin protein for cell culture experiments, dissolve the lyophilized protein in sterile deionized water or 1× PBS at a concentration between 0.1–1.0 mg/mL, depending on your experimental requirements. Avoid vigorous mixing and do not exceed 1 mg/mL to prevent solubility issues.

Step-by-step protocol:

• Preparation:

  • Allow the lyophilized vial to reach room temperature before opening to minimize condensation.
  • Briefly centrifuge the vial to collect all powder at the bottom.

• Reconstitution:

  • Add sterile deionized water or 1× PBS to achieve the desired concentration (commonly 0.1–1.0 mg/mL).
  • Gently mix by pipetting up and down or by slow inversion. Do not vortex or shake vigorously.
  • Let the solution stand at room temperature for 15–30 minutes with gentle agitation to ensure complete dissolution.
  • If solubility is problematic, incubate the solution overnight at 4°C.

• Aliquoting and Storage:

  • For immediate use, keep the reconstituted protein at 4°C for up to 2–7 days.
  • For long-term storage, aliquot and freeze at –20°C or –80°C. Addition of carrier protein (e.g., 0.1% BSA) is recommended to prevent adsorption and loss during freezing.
  • Avoid repeated freeze-thaw cycles.

• Dilution for Cell Culture:

  • Dilute the stock solution into your cell culture medium to the final working concentration required for your experiment.
  • Ensure the final buffer composition is compatible with your cells (e.g., avoid high salt or phosphate concentrations if sensitive).

Additional considerations:

• Confirm protein integrity and concentration after reconstitution by SDS-PAGE or spectrophotometry if needed.
• Check endotoxin levels if using for sensitive cell types; recombinant procalcitonin preparations typically have low endotoxin (<1 EU/µg).
• Always use sterile technique to prevent contamination.

Summary Table:

This protocol ensures optimal solubility, stability, and biological activity for cell culture applications.