A column-switching HPLC-MS/MS method for mucopolysaccharidosis type I analysis in a multiplex assay for the simultaneous newborn screening of six lysosomal storage disorders.

[krabbe disease]

Lysosomal storage disorders comprise a group of rare genetic diseases in which a deficit of specific hydrolases leads to the storage of undegraded substrates in lysosomes . Impaired enzyme activities can be assessed by MS /MS quantification of the reaction products obtained after incubation with specific substrates . In this study , a column-switching HPLC-MS /MS method for multiplex screening in dried blood spot of the lysosomal enzymes activities was developed . Mucopolysaccharidosis type I , Fabry , Gaucher , Krabbe , Niemann-Pick A /B and Pompe diseases were simultaneously assayed . Dried blood spots were incubated with substrates and internal standards ; thereafter , supernatants were collected with minor manipulations . Samples were injected , trapped into an online perfusion column and , by a six -port valve , switched online through the C 18 analytical column to perform separation of metabolites followed by MS /MS analysis . A total of 1136 de -identified newborn screening samples were analyzed to determine references for enzymes activity values . As positive controls , we analyzed dried blood spots from three patients with Pompe , one with Fabry , one with Krabbe disease and two with MPS I , and in all cases the enzyme activities were below the cutoff values measured for newborns , except for an MPS I patient after successful hematopoietic stem cell transplantation .

Diseases
Validation

Diseases presenting "positive controls" symptom

cowden syndrome

erythropoietic protoporphyria

familial hypocalciuric hypercalcemia

krabbe disease

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