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 on line trapping step preceding a liquid chromatography/tandem mass-spectrometry measurement. This method takes only 4 minutes 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.

New Strategy for the Screening of Lysosomal Storage Disorders: The Use of the Online Trapping-and-Cleanup Liquid Chromatography/Mass Spectrometry / G.la Marca; B.Casetta; S. Malvagia; R. Guerrini; E. Zammarchi. - In: ANALYTICAL CHEMISTRY. - ISSN 1520-6882. - ELETTRONICO. - 1;81(15):(2009), pp. 6113-6121.

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

LA MARCA, GIANCARLO;GUERRINI, RENZO;
2009

Abstract

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 on line trapping step preceding a liquid chromatography/tandem mass-spectrometry measurement. This method takes only 4 minutes 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.
1;81(15):
6113
6121
G.la Marca; B.Casetta; S. Malvagia; R. Guerrini; E. Zammarchi
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2158/385854
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