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. 2018 Feb;123(2):135-139.
doi: 10.1016/j.ymgme.2017.10.011. Epub 2017 Oct 23.

Tandem mass spectrometry assay of β-glucocerebrosidase activity in dried blood spots eliminates false positives detected in fluorescence assay

Pavlina Wolf  1 , Roy N Alcalay  2 , Christopher Liong  2 , Emmaline Cullen  3 , Michael W Pauciulo  4 , William C Nichols  4 , Ziv Gan-Or  5 , Wendy K Chung  6 , Tina Faulkner  1 , Christopher Bentis  1 , Robert J Pomponio  1 , Xiwen Ma  1 , X Kate Zhang  1 , Joan M Keutzer  1 , Petra Oliva  7
Affiliations

Tandem mass spectrometry assay of β-glucocerebrosidase activity in dried blood spots eliminates false positives detected in fluorescence assay

Pavlina Wolf et al. Mol Genet Metab. 2018 Feb.

Abstract

Deficiency of β-Glucocerebrosidase (GBA) activity causes Gaucher Disease (GD). GD can be diagnosed by measuring GBA activity (Beutler and Kuhl, 1990). In this study, we assayed dried blood spots from a cohort (n=528) enriched for GBA mutation carriers (n=78) and GD patients (n=18) using both the tandem mass spectrometry (MS/MS) and fluorescence assays and their respective synthetic substrates. The MS/MS assay differentiated normal controls, which included GBA mutation carriers, from GD patients with no overlap. The fluorescence assay did not always differentiate normal controls including GBA mutation carriers from GD patients and false positives were observed. The MS/MS assay improved specificity compared to the fluorescence assay.

Keywords: Dried blood spots (DBS); Gaucher disease (GD); Glucocerebroside; Lysosomal storage disorder (LSD); Newborn screening (NBS); β-Glucocerebrosidase (GBA).

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Figures

Figure 1
Figure 1. Synthetic substrates schema
a. C12-Glucocerebroside used in the MS/MS assay b. 4-methylumbelliferyl β-D-glucopyranoside used in the fluorescence assay.
Figure 2
Figure 2. Fluorescence and mass spectrometry assay correlation
The GBA activity levels in control DBS (normal DBS plus GBA carriers) and GD DBS samples as measured by the MS/MS and the fluorescence (4-MU) assays. The red dotted lines indicates LOD for each assay as described in methods. R=0.8073.
Figure 3
Figure 3. Frequency distribution of GBA enzyme activities in each assay
a. GBA activity distribution in the MS/MS assay. b. GBA activity distribution in the fluorescence assay.
See this image and copyright information in PMC

References

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