A precise, sensitive and stable LC-MSMS method for detection of picomolar levels of serum aldosterone
Background: Accurate quantification of aldosterone is essential in the diagnosis of primary aldosteronism. Liquid chromatography-tandem mass spectrometry (LC-MSMS) analysis is increasingly being used to improve analytical sensitivity and specificity, since this technology reduces most of the interferences observed with immunological methods.
Methods: Serum samples with d7-aldosterone as internal standard were extracted with methyl tert-butyl ether, using liquid-liquid extraction (LLE). Chromatographic separation was performed on a C18 reverse phase column with a methanol-water gradient containing ammonium fluoride. Aldosterone detection was performed on an Agilent 6490 triple quadrupole using electro spray ionisation in positive mode.
Results: Multiple reaction monitoring transitions were m/z 361.2–315.1 for aldosterone, and 368.5–323.3 for d7-aldosterone. Chromatographic retention time was 2.7 min. The method’s total CVs at aldosterone concentrations of 45.4 and 1080 pmol/L were 7.0% and 4.8%, respectively. The intra-assay CVs at concentrations of 60.0 and 637 pmol/L were 4.0% and 2.6%, respectively. The method’s LOQ and LOD were 10 and 5 pmol/L, respectively, demonstrating an excellent analytical sensitivity. The upper limit of quantification was set to 5000 pmol/L, corresponding to the highest calibrator concentration. The long-term stability of the method was evident from repeated measurements of external control pools from UKNEQAS over a period of about 3 years, showing CVs between 2.0 and 7.0%.
Conclusions: We have described a precise, sensitive and stable LC-MSMS method for the measurement of serum aldosterone. In addition, due to the use of LLE and a short LC-column, the method is simple to perform, with a short chromatographic run time.