title: Direct assay of iduronate-2-sulfatase for Hunter disease using UPLC-tandem mass spectrometry and fluorogenic substrate.
abstract: OBJECTIVE
  We devised iduronate-2-sulfatase (IDS) enzyme activity assays by combining fluorometric substrate and LC-MS/MS based detection.
  
  
  METHODS
  4-Methylumbelliferyl α-L-idopyranosiduronic acid 2-sulfate (IDS-S) was used as a substrate for IDS. Its enzymatic product, 4-methylumbelliferyl α-L-idopyranosiduronic acid (IDS-P) and internal standard, 4-methylumbelliferyl α-L-idopyranoside (IDS-IS), were directly measured by UPLC-MS/MS. We determined the precision of our enzyme assay and the effects of sample amounts and incubation time based on the results. Dried blood spots (DBSs) of 110 normal newborns and three patients with Hunter disease were analyzed.
  
  
  RESULTS
  IDS-IS, IDS-P and IDS-S were fully separated using UPLC without any ion suppressions. The intra- and inter-assay precisions were 8.5-10.5% and 11.9-15.3%, respectively. The amount of product obtained was proportional to the number of DBSs and increased linearly with the incubation period from 0 to 15 h. The enzyme activities in DBSs from three patients with MPS II were markedly lower than those in the DBSs of 110 normal newborns.
  
  
  CONCLUSIONS
  To the best of our knowledge, this is the first report describing the use of LC-MS/MS for the diagnosis of Hunter disease with a commercially available substrate. Our method would be a rapid and effective screening tool for the diagnosis of Hunter disease with further study.
authors: Department of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea; Department of Laboratory Medicine, Seoul National University Bundang Hospital, Gyeonggi-do, Republic of Korea.
authors: Kyunghoon
authors: K
authors: Lee
authors: Department of Laboratory Medicine, Seoul National University Bundang Hospital, Gyeonggi-do, Republic of Korea.
authors: Sun-Hee
authors: SH
authors: Jun
authors: Department of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea; Department of Laboratory Medicine, Seoul National University Hospital, Seoul, Republic of Korea.
authors: Sang Hoon
authors: SH
authors: Song
authors: Department of Laboratory Medicine & Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea.
authors: Hyung-Doo
authors: HD
authors: Park
authors: Department of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea; Department of Laboratory Medicine, Seoul National University Bundang Hospital, Gyeonggi-do, Republic of Korea.
authors: Kyoung Un
authors: KU
authors: Park
authors: Department of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea; Department of Laboratory Medicine, Seoul National University Bundang Hospital, Gyeonggi-do, Republic of Korea. Electronic address: songjhcp@snu.ac.kr.
authors: Junghan
authors: J
authors: Song
mesh_terms: D001681:Biological Assay; D015415:Biomarkers / Q000097:blood; D002851:Chromatography, High Pressure Liquid; D059788:Dried Blood Spot Testing / Q000295:instrumentation; D015870:Gene Expression; D006023:Glycoproteins / Q000097:blood* / Q000235:genetics; D006801:Humans; D007067:Iduronic Acid / Q000031:analogs & derivatives; D007231:Infant, Newborn; D008759:Methylglycosides / Q000737:chemistry; D016532:Mucopolysaccharidosis II / Q000097:blood* / Q000175:diagnosis / Q000235:genetics; D015997:Neonatal Screening / Q000379:methods*; D053719:Tandem Mass Spectrometry
chemical_list: D015415:Biomarkers; D006023:Glycoproteins; C018322:IDS protein, human; D008759:Methylglycosides; D007067:Iduronic Acid
keywords: Dried blood spot; Hunter disease; Mucopolysaccharidosis; Newborn screening; Tandem mass spectrometry