title: Digital microfluidic platform for multiplexing enzyme assays: implications for lysosomal storage disease screening in newborns.
abstract: BACKGROUND
  Newborn screening for lysosomal storage diseases (LSDs) has been gaining considerable interest owing to the availability of enzyme replacement therapies. We present a digital microfluidic platform to perform rapid, multiplexed enzymatic analysis of acid α-glucosidase (GAA) and acid α-galactosidase to screen for Pompe and Fabry disorders. The results were compared with those obtained using standard fluorometric methods.
  
  
  METHODS
  We performed bench-based, fluorometric enzymatic analysis on 60 deidentified newborn dried blood spots (DBSs), plus 10 Pompe-affected and 11 Fabry-affected samples, at Duke Biochemical Genetics Laboratory using a 3-mm punch for each assay and an incubation time of 20 h. We used a digital microfluidic platform to automate fluorometric enzymatic assays at Advanced Liquid Logic Inc. using extract from a single punch for both assays, with an incubation time of 6 h. Assays were also performed with an incubation time of 1 h.
  
  
  RESULTS
  Assay results were generally comparable, although mean enzymatic activity for GAA using microfluidics was approximately 3 times higher than that obtained using bench-based methods, which could be attributed to higher substrate concentration. Clear separation was observed between the normal and affected samples at both 6- and 1-h incubation times using digital microfluidics.
  
  
  CONCLUSIONS
  A digital microfluidic platform compared favorably with a clinical reference laboratory to perform enzymatic analysis in DBSs for Pompe and Fabry disorders. This platform presents a new technology for a newborn screening laboratory to screen LSDs by fully automating all the liquid-handling operations in an inexpensive system, providing rapid results.
authors.affiliation: Advanced Liquid Logic, Research Triangle Park, NC, USA.
authors.forename: Ramakrishna S
authors.initials: RS
authors.lastname: Sista
authors.forename: Allen E
authors.initials: AE
authors.lastname: Eckhardt
authors.forename: Tong
authors.initials: T
authors.lastname: Wang
authors.forename: Carrie
authors.initials: C
authors.lastname: Graham
authors.forename: Jeremy L
authors.initials: JL
authors.lastname: Rouse
authors.forename: Scott M
authors.initials: SM
authors.lastname: Norton
authors.forename: Vijay
authors.initials: V
authors.lastname: Srinivasan
authors.forename: Michael G
authors.initials: MG
authors.lastname: Pollack
authors.forename: Adviye A
authors.initials: AA
authors.lastname: Tolun
authors.forename: Deeksha
authors.initials: D
authors.lastname: Bali
authors.forename: David S
authors.initials: DS
authors.lastname: Millington
authors.forename: Vamsee K
authors.initials: VK
authors.lastname: Pamula
mesh_terms: D004796:Clinical Enzyme Tests / Q000295:instrumentation*; D000795:Fabry Disease / Q000175:diagnosis*; D005470:Fluorometry; D006009:Glycogen Storage Disease Type II / Q000175:diagnosis*; D006801:Humans; D007231:Infant, Newborn; D056656:Lab-On-A-Chip Devices; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D015997:Neonatal Screening; D000519:alpha-Galactosidase / Q000097:blood*; D000520:alpha-Glucosidases / Q000097:blood*
chemical_list: D000520:alpha-Glucosidases; D000519:alpha-Galactosidase