Tandem Mass Spectrometry for the Direct Assay of Lysosomal Enzymes in Dried Blood Spots: Application to Screening Newborns for Mucopolysaccharidosis I

BACKGROUND: Treatments now available for mucopolysaccharidosis I require early detection for optimum therapy. Therefore, we have developed an assay appropriate for newborn screening of the activity of the relevant enzyme, alpha-L-iduronidase. METHODS: We synthesized a new alpha-L-iduronidase substrate that can be used to assay the enzyme by use of tandem mass spectrometry together with an internal standard or by fluorometry. The assay uses a dried blood spot on a newborn screening card as the enzyme source. The assay protocol uses a simple liquid-liquid extraction step before mass spectrometry. We optimized enzyme reaction conditions and procedures for the assay, including the concentration of substrate, the reaction pH, the incubation time, and mass spectrometer operation. We also assessed inter- and intraassay imprecision. RESULTS: When the assay was tested on dried blood spots, the alpha-L-iduronidase activity measured for 5 patients with mucopolysaccharidosis I was well below the interval found for 10 randomly chosen newborns. Inter- and intraassay imprecision were <10%. The synthesis of the alpha-L-iduronidase substrate is practical for use on a scale needed to Support newborn screening demands. CONCLUSIONS: This newly developed tandem mass spectrometry assay has the potential to be adopted for newborn screening of mucopolysaccharidosis I. This assay has advantages over a previously reported assay also developed in this laboratory and has the potential to be performed in a multiplex fashion to measure several lysosomal enzymes relevant to treatable lysosomal storage diseases.