Metrological Traceability of the Waters™ MassTrak™ Steroid Serum Set 2 and 3 Calibrators and Quality Controls

Significance of the topic

Liquid chromatography–tandem mass spectrometry (LC-MS/MS) has become the preferred technique for steroid hormone analysis in clinical laboratories, offering superior sensitivity, selectivity, and robustness compared with immunoassays. Regulatory frameworks such as ISO 15189:2022 require clinical laboratories to use metrologically traceable calibration materials to ensure consistent and accurate patient results.

Objectives and study overview

This application note summarizes the design, value assignment, and performance evaluation of Waters MassTrak Steroid Serum Calibrator and Quality Control Sets 2 and 3. It highlights methodologies used to establish metrological traceability, compares assigned values against reference laboratories, and assesses lot-to-lot accuracy and measurement uncertainty.

Metrological traceability methodology

Traceability is achieved via a two-tier approach:

• Primary in-house standards: NIST-traceable reference materials for androstenedione, DHEAS, 11-deoxycortisol, and 21-deoxycortisol, gravimetrically prepared and value assigned.
• Secondary in-house standards: Reference measurement procedure–assigned values for testosterone, cortisol, and 17-hydroxyprogesterone, traceable to University of Ghent and Referenzinstitut für Bioanalytik.

These standards form the basis for assigning concentrations to lyophilized serum calibrators and QCs, with further confirmation via external quality assessment (EQA) schemes and independent in-house panels.

Used Instrumentation

Calibration and validation were performed using a Waters UPLC system coupled to a triple quadrupole mass spectrometer configured for multiple reaction monitoring of steroid analytes. The workflow adheres to established LC-MS/MS protocols for clinical biochemistry.

Main results and discussion

Value assignments of secondary standards showed agreement with reference laboratories within ±6.4% across analytes, with deviations for cortisol, progesterone (demonstration analyte), and testosterone below 4%. Lot-to-lot comparisons for Sets 2 and 3 demonstrated accuracy within ±10% of target across low, medium, and high concentration levels for key steroids. Expanded measurement uncertainty calculations incorporated precision, accuracy, material uncertainty, and instrumentation variability, confirming robust performance.

Benefits and practical applications

• Compliance with ISO 15189:2022 through fully traceable calibrators and QCs.
• High confidence in accuracy and reproducibility of steroid measurements across batches and laboratories.
• Lyophilized format reduces preparation time and enhances stability.

Future trends and potential applications

Advances may include extension of traceable calibrator sets to additional steroid panels, integration with automated sample-preparation platforms, and adoption of digital certification for real-time traceability. Harmonization efforts across global reference laboratories will further standardize clinical steroid assays.

Conclusion

Waters MassTrak Steroid Serum Calibrator and Quality Control Sets 2 and 3 provide fully metrologically traceable materials that meet regulatory requirements and support accurate, reproducible LC-MS/MS steroid analysis in clinical laboratories.

References

• Foley D, Rossiter P, Breen N, Calton LJ. Metrological Traceability of the Waters MassTrak Endocrine Steroid Calibrator and Quality Control Sets. Waters Application Note 720007404. 2022.
• Pitt J. Principles and Applications of Liquid Chromatography–Mass Spectrometry in Clinical Biochemistry. Clinical Biochemist Reviews. 2009 Feb;30(1):19–34.