Abstract
Plasma renin activity (PRA) is an essential analytical tool for screening and diagnosis of secondary forms of hypertension. Typically, PRA is measured by competitive radioimmunoassay, but there are significant drawbacks to this technique including non-specificity, long analysis times, narrow calibration range, and the requirement for radionucleotides. In this paper, we report a method for plasma renin activity determination by immuno-MALDI mass spectrometry detection. This method overcomes the issues of non-specificity and long analytical times present with RIA, and does not require the use of radionucleotides. As an initial methodological evaluation, plasma renin activity results obtained by radioimmunoassay, LC/ESI-MS/MS, and immuno-MALDI on 64 samples from an outpatient primary aldosteronism screening program have been compared. A strong correlation was found between immuno-MALDI and radioimmunoassay (R2 = 0.9412, 62/64 within the 95% CI of the Bland-Altman plot), and iMALDI and LC/ESI-MS/MS (R2 = 0.9471, 62/64 within the 95% CI of the Bland-Altman plot). Technical replicates showed a 4.8% CV, while inter- and intra-day replicates showed CVs of 17.3% and 17.2% respectively. We have developed an assay capable of measuring PRA without the use of radionucleotides. This immuno-MALDI approach affords the specificity of MS while avoiding the long analytical run times and technical problems associated with HPLC. With the use of robotic sample preparation to optimize precision, this assay should be adaptable to clinical environments.
Highlights
Hypertension is a world-wide epidemic affecting more than 1 billion people and causing 7.1 million deaths per year [1]
In the immunocapture coupled to MALDI analysis (iMALDI) method, the amount of angiotensin present is determined by direct MALDI analysis of the affinity beads which are placed on the MALDI target without prior elution of the captured analyte
These two methods were used as comparison methods to judge the correlation of the iMALDI results with those from a Plasma renin activity (PRA) assay at pH 7.4 and pH 6.0
Summary
Hypertension is a world-wide epidemic affecting more than 1 billion people and causing 7.1 million deaths per year [1]. There has been movement away from traditional PRA assays because they are not readily amenable to automation In their place, sandwich assays measuring plasma renin concentration (PRC) have become commonplace, since the development of automated chemiluminescent approaches [11,12,13]. Published MS-based methods have used solidphase extraction (SPE) and positive ion LC/ESI-MS/ MS [21,22] This approach is radionucleotidefree, LC/ESI-MS/MS requires considerable expertise and many clinical laboratories have shied away from this technique because of its technical demands [23]. We have developed a PRA assay using a MALDI platform (Figure 1) This assay uses immunocapture coupled to MALDI analysis (iMALDI) [27,28,29,30], is free of radionucleotides, does not require HPLC, and shows good correlation with existing clinical RIA and LC/ESI-MS/MS methods
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