Abstract:

Objective To establish a candidate reference measurement procedure for serum total homocysteine （Hcy） by liquid chromatography-tandem mass spectrometry （LC-MS/MS）,and to evaluate its analytical performance. Methods Samples were prepared by a simple protein precipitation method,and LC-MS/MS was used to determine serum total Hcy quantitatively. According to the Clinical and Laboratory Standards Institute（CLSI） C62-A and C50-A documents,the performance,including linearity,limit of detection,limit of quantitation,relative matrix effect,precision and trueness,was evaluated. Results The linear range of total Hcy by LC-MS/MS was 0.5-200.0 μmol/L. The limit of quantitation and limit of detection were 0.31 nmol/g and 0.06 nmol/g,respectively. The relative matrix effects were 1.94%,1.91% and 1.78% for sera and solution mixtures in 3 different ratios（1∶1,80∶20 and 20∶80）. The within-run and between-run coefficients of variation（CV）were <2% and <1%. The average recovery rates of 3 levels （30.58,49.21 and 65.42 nmol/g） were 99.8%,100.2%,100.8%, respectively. The bias of NIST standard reference material （SRM） 1950 was <1%. After the samples were processed,they were placed at room temperature [（23±2）℃] and autosampler（10℃） for 24 h, and the results were stable. Conclusions The candidate reference measurement procedure for serum total Hcy by LC-MS/MS has been established,and it has good accuracy and precision,which can be used for measurement traceability.

Key words: Total homocysteine, Isotope-dilution liquid chromatography-tandem mass spectrometry, Reference measurement procedure, Performance evaluation