Homocysteine measurement by Vitros Microtip homocysteine assay.

Background: Elevated circulating total homocysteine is an independent vascular risk factor. Enzymatic homocysteine measurements represent an alternative to HPLC- or immunochemistry-based assays, suitable for automation. Here, we report on analytical performance of a commercial cystathionine β-synthase-based assay, for use on Vitros® automated analyzers. Methods: Linear range, limit of detection and analytical sensitivity were inferred from duplicate measurements of homocystine standard solutions (1-65 μmol/L). Imprecision was assessed using commercial controls according to NCCLS EP5-A2 and accuracy using NIST-SRM1955 reference material. Agreement with a clinically validated HPLC method was examined on 207 patient samples. Results: The enzymatic assay was linear from 1 to 90 μmol/L homocysteine. Total (within-day) imprecision ranged from 4.5 (3.9)% to 2.8 (1.6)% at homocysteine 9.7-43.2 μmol/L. Accuracy was acceptable at 8.9 and 17.7 μmol/L homocysteine, with +6.4% and -1.2% bias, respectively, but showed substantial negative bias (-20.1%) at 4.0 μmol/L. High triglycerides (19.8 μmol/L) negatively interfered. The enzymatic method was slightly less sensitive than the HPLC method (limit of detection 0.7 and 0.2 μmol/L, respectively) but correlated well with the latter (r 2 =0.9997, slope=1.04, intercept=-0.66 μmol/L) and was more precise (p<0.05). Conclusions: The Vitros® homocysteine assay met the CLIA Desirable Analytical Quality Specifications at homocysteine ≥9 μmol/L. Its analytical performance and suitability for automation make the Vitros® assay an analytically acceptable alternative to HPLC-based methods.