Total testosterone quantitative measurement in serum by LC-MS/MS

Area of Science:

• Clinical Chemistry
• Analytical Chemistry
• Biochemistry

Background:

• Accurate total testosterone measurement is crucial for diagnosing and managing various endocrine disorders in adults and children.
• Existing methods may have limitations in precision or accuracy, necessitating the development of improved analytical techniques.

Purpose of the Study:

• To develop and validate a precise and accurate mass spectrometric method for quantifying total testosterone in human serum.
• To establish a reliable assay suitable for routine clinical diagnostics and research.

Main Methods:

• A novel method utilizing liquid chromatography-isotopic dilution tandem mass spectrometry (LC-ID-MS/MS) for total testosterone determination.
• Sample preparation involves automated protein release, dual liquid-liquid extraction steps to isolate testosterone, and reconstitution in a basic buffer.
• Quantification is performed using a liquid-handling system with 96-deepwell plates for high-throughput processing.

Main Results:

• The developed method demonstrates a low limit of detection (9.71 pmol/L or 0.280 ng/dL).
• Average percent bias was not significantly different from established reference methods, indicating high accuracy.
• Excellent long-term precision was observed over two years, with coefficients of variation ranging from 3.7% to 4.8% across different concentration levels.

Conclusions:

• This validated LC-ID-MS/MS method offers consistently high accuracy and excellent precision for total testosterone measurement in human serum.
• The method is suitable for clinical applications across all relevant concentration ranges, supporting reliable diagnosis and monitoring of hormonal conditions.