Analysis of glucocerebrosidase activity in dry blood spots using tandem mass spectrometry

Background: Gaucher disease (GD) is due to deficiency of acid-β-glucosidase (ABG) and comprises a clinical spectrum with variable age of onset and severity. We evaluated a tandem mass spectrometry (MS/MS) method to measure ABG activity for high through-put screening. Methods: ABG activity was measured in 3.2mm punches from dry blood spots (DBS). Each punch was incubated for 21h with the substrate D-Glucosyl-β1-1β-N-dodecanoyl-D-erythro-sphingosine [C12-glucocerebroside (C₃₆H₆₉NO₈)] and internal standard N-myristoyl-D-erythro-sphingosine [C14-ceramide (C₃₂H₆₃NO₃)]. The product and internal standard were quantified using MS/MS. Results: ABG activities in anonymized newborn screening samples from NY State were (mean) 22.0 μmol/h/L ± (SD) 13.8 μmol/h/L (n=2088, median 19.9 μmol/h/L, 95%CI 22.59-21.41 μmol/h/L). The enzymatic activity in DBS from 10 treatment naïve adult Gaucher patients was less than 4.2 μmol/h/L. ABG activity was stable for 3. months at room temperature a 20% activity reduction was observed. Inter- and intra-run imprecisions were 8% and 13.7%, respectively. The limit of detection was 0.75 μmol/h/L and limit of quantification was 1.25 μmol/h/L. Conclusions: The measurement of ABG activities in DBS using MS/MS is suitable for high-throughput analysis of at-risk individuals and potentially for newborn screening for GD.