Validation of a Multiplex Tandem Mass Spectrometry Method for the Detection of Selected Lysosomal Storage Diseases in Dried Blood Spots

Abstract

Background: Interest in screening methods for lysosomal storage diseases (LSDs) has increased in recent years, since early diagnosis and treatment are essential to prevent or attenuate the onset of symptoms and the complications of these diseases. In the current work, we evaluated the performance of tandem mass spectrometry (MS/MS) for the detection of some LSDs, aiming the future use of this methodology for the screening of these disorders. Methods: Standard curves and quality control dried blood spots were assayed to evaluate the precision, linearity, and accuracy. A total of 150 controls were grouped according to age and subjected to measurement of lysosomal enzymes deficient in Niemann-Pick A/B, Krabbe, Gaucher, Fabry, Pompe, and Mucopolysaccharidosis type I diseases. Samples from 59 affected patients with a diagnosis of LSDs previously confirmed by fluorimetric methods were analyzed. Results: Data from standard calibration demonstrated good linearity and accuracy and the intra-and interassay precisions varied from 1.17% to 11.60% and 5.39% to 31.24%, respectively. Except for galactocerebrosidase and a-L-iduronidase, enzyme activities were significantly higher in newborns compared to children and adult controls. Affected patients presented enzymatic activities significantly lower compared to all control participants. Conclusion: Our results show that MS/MS is a promising methodology, suitable for the screening of LSDs, but accurate diagnoses will depend on its correlation with other biochemical and/or molecular analyses.

Reduced α-galactosidase A activity in zebrafish (Danio rerio) mirrors distinct features of Fabry nephropathy phenotype

“…Most interestingly, the human α-GAL activity assays can effectively be used to measure zebrafish enzyme activity and we could therefore demonstrate that it is present in zebrafish as early as in 2 dpf embryo, 4 dpf larva, 30 dpf juvenile and adult fish tissues 90 + dpf ( Fig. 2 B), and fluctuates similarly to humans [56] . We have noticed that enzyme activity in general is higher in adult wildtype female compared to adult male zebrafish, in agree with what have been described previously in Fabry mice [57] as well as in human [58] .…”

Application of MALDI-TOF Mass Spectrometry for Non-invasive Diagnostics of Mucopolysaccharidosis IIIA

“…Some LSDs were detected by measuring of activities of their corresponding enzymes (galactocerebroside-βgalactosidase for the analysis of Krabbe disease; α-galactosidase A for the analysis of Fabry disease) in dried blood spots using tandem mass spectrometry (MS/MS) [10]. MS/MS analysis of dried blood spots is also useful for enzymatic-based newborn screening of every MPS type except MPS IX [11][12][13]. MS analysis in combination with liquid chromatography (LC-MS) was used for GAG detection in urine and blood samples [14][15][16].…”

Development of a fast LC-MS/MS protocol for combined measurement of six LSDs on dried blood spot in a newborn screening program

“…Many protocols for lysosomal enzyme activation and measurement have been developed to allow the simultaneous detection of several enzymes on one or two DBS punches [12][13][14][15].…”