Reliable in-house LC-MS/MS method for steroid hormone analysis: Validations and comparisons.

Area of Science:

• Clinical Chemistry
• Analytical Chemistry
• Endocrinology

Background:

• Accurate steroid hormone quantification is vital for diagnosing and monitoring endocrine disorders.
• Liquid chromatography-tandem mass spectrometry (LC-MS/MS) provides high specificity and sensitivity for analyzing multiple steroids simultaneously.

Purpose of the Study:

• To develop and validate a high-throughput LC-MS/MS method for quantifying 19 steroids in human plasma and serum.
• To compare the developed method's performance against a routine chemiluminescence immunoassay and a commercial LC-MS/MS assay.

Main Methods:

• A high-throughput LC-MS/MS method was developed for 19 steroids using optimized protein precipitation and solid-phase extraction.
• The method was validated according to established guidelines.
• Performance was compared with an immunoassay (n=5 steroids) and a commercial LC-MS/MS method (n=17 steroids) using 208 plasma samples.

Main Results:

• The developed LC-MS/MS method showed excellent linearity (R² > 0.992), sensitivity (LOD: 0.05-0.5 ng/mL), precision, and accuracy (CV < 15%, recovery: 91.8%-110.7%).
• Good correlation was observed with immunoassay (ICCs > 0.90), with superior accuracy at low concentrations for testosterone and progesterone.
• High concordance (ICCs > 0.96) and minimal bias were found compared to a commercial LC-MS/MS method.

Conclusions:

• The developed LC-MS/MS method is reliable and suitable for clinical laboratory applications.
• It provides a more accurate alternative to immunoassays for steroid hormone analysis.
• Strong agreement with a validated commercial LC-MS/MS assay confirms its robustness and clinical utility.