New Strategy for the Screening of Lysosomal Storage Disorders: The Use of the Online Trapping-and-Cleanup Liquid Chromatography/Mass Spectrometry

被引:49
作者
la Marca, Giancarlo [1 ,2 ]
Casetta, Bruno [3 ]
Malvagia, Sabrina [4 ]
Guerrini, Renzo [4 ]
Zammarchi, Enrico [5 ]
机构
[1] Univ Florence, Dept Pharmacol, Mass Spectrometry & Pharmacol Lab, A Meyer Childrens Hosp,Pediat Neurol Unit, Florence, Italy
[2] Univ Florence, Dept Pharmacol, Mass Spectrometry & Pharmacol Lab, Pediat Neurol Labs,A Meyer Childrens Hosp, Florence, Italy
[3] Appl Biosyst Inc, I-20052 Monza, Italy
[4] Univ Florence, Dept Neurosci, Florence, Italy
[5] Univ Florence, Dept Pediat, I-50139 Florence, Italy
关键词
ENZYME REPLACEMENT THERAPY; DRIED BLOOD SPOTS; TANDEM MASS-SPECTROMETRY; ALPHA-GLUCOSIDASE ACTIVITY; DIRECT MULTIPLEX ASSAY; FABRY-DISEASE; MUCOPOLYSACCHARIDOSIS-VI; CELL TRANSPLANTATION; AGALSIDASE-ALPHA; DIAGNOSIS;
D O I
10.1021/ac900504s
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
The aim of this study was to set up a robust method suitable for large-scale studies (screening) with a minimized preparation process and with reduced running costs, for measuring five enzyme activities on dried blood spots by a new and simplified tandem mass spectrometry-based method. After incubation, all 5 reaction mixtures, carried out separately, were stopped, combined together, and centrifuged. The cleaning-up of the injected mixture was performed through a fast online trapping step preceding a liquid chromatography/tandem mass-spectrometry measurement. This method takes only 4 min as analysis run time and without any purification following the enzymatic reaction. We assessed the effectiveness of this approach in assaying the enzymatic activities on dried blood spots from 10 patients affected by "Pompe", 6 by "Gaucher", 12 by "Fabry", 3 by "Niemann-Pick" A/B, and 2 by "Krabbe" diseases. Reference values were established on 5000 healthy newborns and 300 healthy adults. All affected patients showed enzymatic activities below the normal range. In heterozygous carriers (18 for Fabry, 10 for Pompe, and 4 for Gaucher disease) the activities were slightly lower than in control subjects. The results show that the method set out in its simplicity, low costs, and low processes preparations can be fully applicable to a mass screening.
引用
收藏
页码:6113 / 6121
页数:9
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