BACKGROUND: Pre-symptomatic hematopoietic stem cell transplantation is essential to achieve best possible outcomes for patients with the childhood cerebral form of X-linked adrenoleukodystrophy (X-ALD). We describe a high-throughput method for measurement of C20-C26 lysophosphatidylcholines (LPCs) and biochemical diagnosis of X-ALD using the same dried blood spots (DBS) routinely used for newborn screening. METHODS: LPCs are extracted from 3-mm DBS punch with methanol containing an isotopically labeled LPC as internal standard. This extract is transferred to a 96-well plate, evaporated and then reconstituted in mobile phase for flow injection analysis tandem mass spectrometry (FIA-MS/MS) in selected reaction monitoring mode for measurement of four different LPCs (C20, C22, C24, C26) and the internal standard (d4-C26-LPC). Analysis time is 1.5min per sample. RESULTS: The mean CVs from the intra- and inter-assay experiments for LPCs were 6.3-15.1% for C20-LPC, 4.4-18.6% for C22-LPC and 4.5-14.3% for C24-LPC. Limits of detection were determined for C20-LPC (LOD=0.03μg/mL), C22-LPC (0.03μg/mL), C24-LPC (0.03μg/mL) and C26-LPC (0.01μg/mL). Reference ranges were established from DBS of 130 newborns and 20 adults. Samples of patients with X-ALD (n=16), peroxisomal biogenesis disorders (n=8), and X-ALD carriers (n=12) were analyzed blindly and all were correctly identified. CONCLUSION: Analysis of LPC species by FIA-MS/MS is a fast, simple and reliable method to screen for X-ALD and other peroxisomal disorders in DBS. To maximize specificity, abnormal results can be verified by a 2nd tier assay using LC-MS/MS.
BACKGROUND: Pre-symptomatic hematopoietic stem cell transplantation is essential to achieve best possible outcomes for patients with the childhood cerebral form of X-linked adrenoleukodystrophy (X-ALD). We describe a high-throughput method for measurement of C20-C26 lysophosphatidylcholines (LPCs) and biochemical diagnosis of X-ALD using the same dried blood spots (DBS) routinely used for newborn screening. METHODS:LPCs are extracted from 3-mm DBS punch with methanol containing an isotopically labeled LPC as internal standard. This extract is transferred to a 96-well plate, evaporated and then reconstituted in mobile phase for flow injection analysis tandem mass spectrometry (FIA-MS/MS) in selected reaction monitoring mode for measurement of four different LPCs (C20, C22, C24, C26) and the internal standard (d4-C26-LPC). Analysis time is 1.5min per sample. RESULTS: The mean CVs from the intra- and inter-assay experiments for LPCs were 6.3-15.1% for C20-LPC, 4.4-18.6% for C22-LPC and 4.5-14.3% for C24-LPC. Limits of detection were determined for C20-LPC (LOD=0.03μg/mL), C22-LPC (0.03μg/mL), C24-LPC (0.03μg/mL) and C26-LPC (0.01μg/mL). Reference ranges were established from DBS of 130 newborns and 20 adults. Samples of patients with X-ALD (n=16), peroxisomal biogenesis disorders (n=8), and X-ALD carriers (n=12) were analyzed blindly and all were correctly identified. CONCLUSION: Analysis of LPC species by FIA-MS/MS is a fast, simple and reliable method to screen for X-ALD and other peroxisomal disorders in DBS. To maximize specificity, abnormal results can be verified by a 2nd tier assay using LC-MS/MS.
Authors: Thais Armangue; Joseph J Orsini; Asako Takanohashi; Francesco Gavazzi; Alex Conant; Nicole Ulrick; Mark A Morrissey; Norah Nahhas; Guy Helman; Heather Gordish-Dressman; Simona Orcesi; Davide Tonduti; Chloe Stutterd; Keith van Haren; Camilo Toro; Alejandro D Iglesias; Marjo S van der Knaap; Raphaela Goldbach Mansky; Anne B Moser; Richard O Jones; Adeline Vanderver Journal: Mol Genet Metab Date: 2017-07-20 Impact factor: 4.797
Authors: Stephan Kemp; Irene C Huffnagel; Gabor E Linthorst; Ronald J Wanders; Marc Engelen Journal: Nat Rev Endocrinol Date: 2016-06-17 Impact factor: 43.330
Authors: Eric J Mallack; Bela R Turk; Helena Yan; Carrie Price; Michelle Demetres; Ann B Moser; Catherine Becker; Kim Hollandsworth; Laura Adang; Adeline Vanderver; Keith Van Haren; Maura Ruzhnikov; Joanne Kurtzberg; Gustavo Maegawa; Paul J Orchard; Troy C Lund; Gerald V Raymond; Molly Regelmann; Joseph J Orsini; Elisa Seeger; Stephan Kemp; Florian Eichler; Ali Fatemi Journal: J Inherit Metab Dis Date: 2021-01-09 Impact factor: 4.982
Authors: Andrea Coppa; Sanjib Guha; Stéphane Fourcade; Janani Parameswaran; Montserrat Ruiz; Ann B Moser; Agatha Schlüter; Michael P Murphy; Jose Miguel Lizcano; Antonio Miranda-Vizuete; Esther Dalfó; Aurora Pujol Journal: Free Radic Biol Med Date: 2020-02-01 Impact factor: 7.376
Authors: Malu-Clair van de Beek; Inge M E Dijkstra; Henk van Lenthe; Rob Ofman; Dalia Goldhaber-Pasillas; Nicolas Schauer; Martin Schackmann; Joo-Yeon Engelen-Lee; Frédéric M Vaz; Wim Kulik; Ronald J A Wanders; Marc Engelen; Stephan Kemp Journal: PLoS One Date: 2016-04-28 Impact factor: 3.240