Comparison of isotopic and immunoenzymatic methods for folate and vitamin B12 determination.

BACKGROUND: The objective of this study was to compare automated immunoenzymatic assay (IEA) with an isotopic reference method for determination of plasma folate, erythrocyte folate, and plasma vitamin B12 levels. METHODS: Analytical performance including within-run and between-run imprecision was assessed on the automated IEA on the Access2 (Beckman Coulter, France). Folate and B12 levels determined routinely with a radioimmunoassay (RIA) (Simultrac, Diasorin, Italy) using EDTA samples (n=100) were compared to the Access2 IEA. To evaluate pre-analytical conditions, EDTA and heparinized plasma (n=56) from simultaneous samplings were used to compare folate and B12 levels via RIA and the Access2. In addition, discordant results were assayed by IEA on the Architect i2000 (Abbott, France). RESULTS: Coefficient of variation for folate and B12 measurements on the Access2 ranged between 1.5% and 4.1%, respectively, for within-run assays, while between-run imprecision ranged from 4.8% to 9.3%, respectively. Comparison of RIA (x) and IEA on the Access2 (y) using EDTA samples showed a good correlation for plasma folate (y = 0.94x + 1.09; r2 = 0.83), acceptable correlation for erythrocyte folate (y = 0.65x + 137; r2 = 0.57) and discrepancy for B12 (y = 0.67x + 211; r2 = 0.37). This discordance for vitamin B12 was not observed when comparing RIA (x) and IEA on the Architect i2000 (y) (y = 0.74x + 40; r2 = 0.92) and disappeared when using heparinized samples for the Access2 (y = 0.85x + 7.05; r2 = 0.90). CONCLUSIONS: Automated IEA appears to be a good alternative to RIA for the determination of folate and vitamin B12 but requires the use of an adequate anticoagulant.

A New DiaSys colorimetric assay for plasma homocysteine: application in diabetic patients.

This study was conducted to evaluate the analytical performance of a new colorimetric assay for plasma homocysteine (Hcy) and to determine the influence of the method used on the frequency of hyperhomocysteinemia and on the association between Hcy and biological parameters in diabetic patients. Plasma Hcy was measured in 160 diabetic patients by high performance liquid chromatography (HPLC) in parallel with an automated colorimetric assay (DiaSys Diagnostic Systems GmbH) using the Olympus AU640 and Abbott C8000 analyzers. Plasma glucose, creatinine, folates, vitamin B(12), high-sensitivity C-reactive protein (hs-CRP), and blood hemoglobin A1c (HbA1c) concentrations were measured by routine methods. The Hcy concentrations obtained with the HPLC (x) and colorimetric methods (y) were highly correlated (y = 1.032x - 0.65, r(2) = 0.884 for AU640; y = 1.028x + 0.87, r(2) = 0.934 for C8000). The frequency of Hcy >15 micromol/L was 41.9% by HPLC assay and there was no statistically significant difference using the colorimetric method. Patients with Hcy >15 micromol/L compared to < or =15 micromol/L using HPLC had significantly lower glycemia (p <0.001), lower HbA1c (p <0.001), lower estimated glomerular filtration rate (GFR) (p <0.0001), and lower vitamin B(12) levels (p <0.001). All these relationships were also noted when using the colorimetric assay on the Olympus AU640 or Abbott C8000 analyzers. In summary, this study shows that the DiaSys colorimetric assay is suitable for plasma Hcy determinations in routine clinical biochemistry.