SFEBES2012 Poster Presentations Steroids (33 abstracts)
Introduction: Aldosterone is a potent adrenal steroid hormone which regulates renal sodium reabsorption and potassium secretion. Clinically, measurement of aldosterone is important to identify patients with primary hyperaldosteronism, and has traditionally been performed by immunoassay. The specificity of immunoassays is often poor due to antibody cross reactivity to other structurally related steroid hormones, resulting in falsely elevated concentrations. To overcome these limitations we have developed a highly sensitive and specific method for the measurement of aldosterone from human plasma by UPLC-MS/MS.
Methods: Calibrators and quality control (QC) material were prepared by spiking aldosterone into phosphate buffered saline containing 0.1% bovine serum albumin. Calibrators, QCs and patient samples (250 µl) were extracted using a Waters Oasis® HLB 96-well extraction plate. Chromatography was performed on a Phenomenex Kinetex 2.6 µm XB-C18 100×3.0 mM column, and LC-MS/MS on a Waters Xevo TQS instrument in electrospray negative ionisation mode.
Results: The assay was linear up to 3200 pmol/L, and the lower limit of quantitation was 25 pmol/L. Ion suppression studies demonstrated minimal ion suppression, defined as a fall or rise in baseline total ion count ≥10% at the retention time of the analyte. Intra and inter-assay imprecision, assessed by repeat measurements of three QC samples covering a clinically relevant concentration range were all ≤10%. Patient plasma samples were used to compare the new UPLC-MS/MS method with an established radioimmunoassay. Passing and Bablock analysis yielded the equation LC-MS/MS = 0.79 (RIA) −41.7, r2 = 0.88, r = 0.94 (n= 54).
Conclusions: We have developed a highly sensitive and specific assay for LC-MS/MS measurement of aldosterone from human plasma. As well as demonstrating improved sensitivity compared to established aldosterone immunoassays in routine clinical use, the UPLC-MS/MS method should also provide a more specific method.
Declaration of interest: There is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.
Funding: No specific grant from any funding agency in the public, commercial or not-for-profit sector.