title: Galactocerebrosidase assay on dried-leukocytes impregnated in filter paper for the detection of Krabbe disease.
abstract: BACKGROUND
  Krabbe disease (KD) is an inherited lysosomal storage disease (LSD) caused by the deficiency of galactocerebrosidase (GALC) and is characterized by a severe and progressive leukodystrophy with death frequently before one year of life in the classical early-onset form. As a consequence of the enzyme defect, globoid cells containing undigested galactosylceramide are observed and are characteristic of the disease. Hematopoietic stem cell transplantation is the current treatment for this disease, with some success in the classical cases if performed very early in life. Definitive diagnosis of KD is generally accessed by determination of GALC in leukocytes or fibroblasts. For the last few years, dried-blood filter paper (DBFP) samples have been increasingly used for lysosomal enzyme assays. Originally, some lysosomal enzymes could not be tested in DBFP samples using fluorometric assays, including GALC, heparan-sulfamidase and a few others. Recently, we reported successful results using dried-leukocytes filter paper (DLFP) samples for heparan sulfamidase and β-galactosidase. Extending these studies, we present now a new GALC assay on these type of samples.
  
  
  METHODS
  Adapted leukocyte fluorometric assay was used for the evaluation of GALC in DLFP samples.
  
  
  RESULTS
  Our results using this method showed a clear discrimination between GALC levels observed in KD patients and healthy controls.
  
  
  CONCLUSIONS
  The assay is robust and reliable and could be adopted by reference laboratories for diagnosis of LSDs. It is expected that the use of DLPF would make it possible to diagnose patients living in isolated areas, where liquid samples usually have to be transported over several days and sometimes across country borders before reaching reference laboratories.
authors.affiliation: Medical Genetics Service, HCPA, Porto Alegre, RS, Brazil; Postgraduate Program in Medicine, Medical Sciences, UFRGS, Porto Alegre, RS, Brazil.
authors.forename: M
authors.initials: M
authors.lastname: Camelier
authors.affiliation: Medical Genetics Service, HCPA, Porto Alegre, RS, Brazil; Department of Genetics, UFRGS, Porto Alegre, RS, Brazil. Electronic address: gecivallero@yahoo.com.
authors.forename: G
authors.initials: G
authors.lastname: Civallero
authors.affiliation: Medical Genetics Service, HCPA, Porto Alegre, RS, Brazil.
authors.forename: J
authors.initials: J
authors.lastname: De Mari
authors.affiliation: Medical Genetics Service, HCPA, Porto Alegre, RS, Brazil.
authors.forename: M
authors.initials: M
authors.lastname: Burin
authors.affiliation: Medical Genetics Service, HCPA, Porto Alegre, RS, Brazil; Postgraduate Program in Medicine, Medical Sciences, UFRGS, Porto Alegre, RS, Brazil; Department of Genetics, UFRGS, Porto Alegre, RS, Brazil; INAGEMP, National Institute of Population Medical Genetics, Porto Alegre, RS, Brazil.
authors.forename: R
authors.initials: R
authors.lastname: Giugliani
mesh_terms: D001681:Biological Assay; D016022:Case-Control Studies; D005698:Galactosylceramidase / Q000378:metabolism*; D006801:Humans; D007962:Leukocytes / Q000201:enzymology*; D007965:Leukodystrophy, Globoid Cell / Q000175:diagnosis* / Q000201:enzymology*; D010209:Paper; D011379:Prognosis
chemical_list: D005698:Galactosylceramidase
keywords: Dried-leukocytes; Enzyme assay; Filter paper samples; Galactocerebrosidase; Krabbe disease; Lysosomal storage diseases