Plasma renin activity (PRA) is essential for the screening and diagnosis of primary aldosteronism (PA), a form of secondary hypertension, which affects approximately 100 million people worldwide. It is commonly determined by radioimmunoassay (RIA) and, more recently, by relatively low-throughput LC-MS/MS methods. In order to circumvent the negative aspects of RIAs (radioisotopes, cross-reactivity) and the low throughput of LC-MS based methods, we have developed a high-throughput immuno-MALDI (iMALDI)-based assay for PRA determination using an Agilent Bravo for automated liquid handling and a Bruker Microflex LRF instrument for MALDI analysis, with the goal of implementing the assay in clinical laboratories. The current assay allows PRA determination of 29 patient samples (192 immuno-captures), within ~6 to 7h, using a 3-hour Ang I generation period, at a 7.5-fold faster analysis time than LC-MS/MS. The assay is performed on 350μL of plasma, and has a linear range from 0.08 to 5.3ng/L/s in the reflector mode, and 0.04 to 5.3ng/L/s in the linear mode. The analytical precision is 2.0 to 9.7% CV in the reflector mode, and 1.5 to 14.3% CV in the linear mode. A method comparison to a clinically employed LC-MS/MS assay for PRA determination showed excellent correlation within the linear range, with an R(2) value of ≥0.98. This automated high throughput iMALDI platform has clinically suitable sensitivity, precision, linear range, and correlation with the standard method for PRA determination. Furthermore, the developed workflow based on the iMALDI technology can be used for the determination of other proteomic biomarkers. This article is part of a Special Issue entitled: Medical Proteomics.
Keywords: Angiotensin-I; Automation; Hypertension; Immuno-MALDI (iMALDI); Matrix assisted laser desorption/ionization (MALDI) mass spectrometry; Plasma renin activity (PRA).
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