Background The current first-line screening test for primary hyperaldosteronism is the plasma aldosterone:renin ratio, however renin assays have several disadvantages and the ARR is affected by medications and physiological factors. Angiotensin II is a key biologically active hormone in the renin-angiotensin-aldosterone system. It has been suggested that measurement of equilibrium levels of this peptide, involving an in vitro incubation of serum prior to analysis, may provide a better marker of renin-angiotensin-aldosterone system activity than renin. Methods An eqAng II LC-MS/MS assay was developed, optimised and validated. Serum samples were incubated at 37 °C for 45 minutes prior to stabilisation with cold EDTA solution, solid phase extraction and LC-MS/MS analysis. Stability in whole blood and the effect of cryoactivation were assessed. For comparison to the current screening test, 150 anonymised patient samples were analysed for eqAng II, renin activity and aldosterone (all by LC-MS/MS). Results The assay had good precision, minimal bias and acceptable recovery. EqAng II did not change significantly when whole blood samples were stored for up to 72 h, and cryoactivation was only observed for pregnant patients. EqAng II was significantly correlated with renin, and the aldosterone:eqAng II ratio had a strong positive correlation with the aldosterone:renin ratio. Conclusions An LC-MS/MS assay for eqAng II has been developed which shows promise as an alternative screening test for primary hyperaldosteronism. Compared to renin assays it is quicker, simpler and less likely to be affected by anti-hypertensive medications. Further clinical validation in hypertensive patients would be required prior to implementation.

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