Analytical and clinical evaluation of the Bayer ADVIA Centaur homocysteine assay.

BACKGROUND: Cardiovascular disease (CVD) is the leading cause of death in the United States. Elevations in homocysteine (Hcy) have been associated with increased risk of acute coronary syndrome, stroke and peripheral vascular disease. Increased utilization of Hcy as a risk marker has prompted the need for high throughput methods that are simple to use and analytically accurate and precise. METHODS: We report the performance characteristics of the automated Bayer ADVIA Centaur chemiluminescent Hcy assay. Centaur Hcy is based on a three-step procedure: (1) reduction of Hcy disulfides to free Hcy, (2) enzymatic conversion of free Hcy to S-adenosyl Hcy (SAH) and (3) quantitation of SAH in a competitive immunoassay (labeled anti-SAH antibody: magnetic particles coupled with SAH). RESULTS: Total assay precision ranged from 3.5% to 6.8% at 4.9-62 micromol/Hcy; linearity undiluted from 0 to 65 micromol/l, up to 650 micromol/l with automatic dilution. Method comparisons with fluorescent polarization immunoassay and high-performance liquid chromatography (HPLC) gave linear regression equations with slopes between 0.95 and 1.0. Measurement of Hcy concentrations in apparently healthy populations yielded middle 95th percentile of 9.7 micromol/l, consistent with epidemiologic studies suggesting that 9-10 micromol/l represents the lower threshold of a population at risk of CVD. CONCLUSIONS: The Centaur Hcy assay is a sensitive and precise assay for the measurement of Hcy.

The effect of class-specific protease inhibitors on the stabilization of B-type natriuretic peptide in human plasma.

BACKGROUND: B-type natriuretic peptide (BNP) is a cardiac hormone that regulates hemodynamic equilibrium. In the circulation, its activity is controlled by proteolytic factors. Accurate measurement of BNP in a patient's plasma may be affected by degradation due to proteolysis. OBJECTIVE: We report on the identification and performance of classes of protease inhibitors that stabilize BNP in plasma. DESIGN AND METHODS: Using the Bayer ADVIA Centaur BNP assay, we measured the effect of arginine, serine and/or specific kallikrein protease inhibitors (PIs) on exogenous spiked or endogenous BNP in patient plasma. RESULTS: Compared to controls without inhibitor, all PIs were capable, to varying degrees, of retarding the rate of proteolytic degradation. The kallikrein-specific inhibitor, D-Phe-Phe-Arg-chloromethylketone (PPACK II) was most effective as a single constituent and was able to eliminate BNP degradation in patient samples for up to 6-10 days when stored at 2-8 degrees C. CONCLUSIONS: The stability of BNP was markedly increased in the presence of kallikrein-specific PPACK II and a broad spectrum of serine PIs. Use of these compounds offers a simple method of extending sample handling and storage of plasma samples containing BNP.