BACKGROUND: Interest in lysosomal storage disorders, a collection of more than 40 inherited metabolic disorders, has increased because of new therapy options such as enzyme replacement, stem cell transplantation, and substrate reduction therapy. We developed a high-throughput protocol that simplifies analytical challenges such as complex sample preparation and potential interference from excess residual substrate associated with previously reported assays.


After overnight incubation (16-20 h) of dried blood spots with a cassette of substrates and deuterated internal standards, we used a TLX-2 system to quantify 6 lysosomal enzyme activities for Fabry, Gaucher, Niemann-Pick A/B, Pompe, Krabbe, and mucopolysaccharidosis I disease. This multiplexed, multidimensional ultra-HPLC-tandem mass spectrometry assay included Cyclone P Turbo Flow and Hypersil Gold C8 columns. The method did not require offline sample preparation such as liquid-liquid and solid-phase extraction, or hazardous reagents such as ethyl acetate.


Obviating the offline sample preparation steps led to substantial savings in analytical time (approximately 70%) and reagent costs (approximately 50%). In a pilot study, lysosomal enzyme activities of 8586 newborns were measured, including 51 positive controls, and the results demonstrated 100% diagnostic sensitivity and high specificity. The results for Krabbe disease were validated with parallel measurements by the New York State Screening Laboratory.


Turboflow online sample cleanup and the use of an additional analytical column enabled the implementation of lysosomal storage disorder testing in a nationwide screening program while keeping the total analysis time to <2 min per sample.

Published in Clinical Chemistry. Read the online article here. 

Authors / corresponding author* 

Thomas F. Metz*,1,2; Thomas P. Mechtler*,1,2;  Joseph J. Orsini, 3;  Monica Martin, 3;  Bori Shushan, 4; Joseph L. Herman, 5; Rene Ratschmann, 1; Chike B. Item, 1,2; Berthold Streubel,6; Kurt R. Herkner,1,2; David C. Kasper,1,2

1 Department of Pediatrics and Adolescent Medicine; Medical University of Vienna
2 Research Core Unit of Pediatric Biochemistry and Analytics, Department of Pediatrics and Adolescent; Medical University of Vienna
3 Biggs Laboratory, Wadsworth Center, New York State Department of Health, Albany, NY
4 Clinical Mass Spec Consultants, Toronto, ON, Canada
5 Thermo Fisher Scientific, 101 Constitution Boulevard, Franklin MA
6 Department of Pathology, Medical University of Vienna, Vienna, Austria