Determination of ACE concentration.

<p>Three patients were selected to represent all genotypes (II, ID and DD, indicated). Sera were diluted 12.5, 25, 50, 100 and 200-fold (indicated) to determine optimal conditions. ACE amount was determined in an ELISA assay. A set samples with known ACE concentrations (Standard, <b>A</b>) were also used. Values (optical density) were plotted as the function of the known ACE concentration and a calibration curve was defined (nonlinear fit, solid line, <b>A</b>). Concentrations of the serum samples were then calculated based on their position on the calibration curve. Each serum sample was determined two times. Average of the quadruple measurements is shown (three samples measured twice, resulting in 6, sometimes overlapping individual data point for each dilution, <b>A</b>). It was apparent from the individual measurements that dilutions at 200-fold have very low values, while the saturation of the binding sites resulted in a high scatter at dilutions below 50-fold (<b>A</b>). Although averaged values showed a tendency for under-estimating ACE concentrations at low dilutions, in the case of high expression levels (<b>B</b>), ACE concentration determination was remarkably accurate. Bars represent mean ± SD. No statistical differences were found by nonparametric ANOVA.