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Mol Genet Metab. 2015 Jan;114(1):46-50. doi: 10.1016/j.ymgme.2014.11.013. Epub 2014 Nov 27.

Streamlined determination of lysophosphatidylcholines in dried blood spots for newborn screening of X-linked adrenoleukodystrophy.

Author information

1
Biochemical Genetics Laboratory, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.
2
Kennedy Krieger Institute, Baltimore, MD 21205, USA.
3
Department of Medical Biochemistry, Oslo University Hospital, Oslo, Norway.
4
Biochemical Genetics Laboratory, Mayo Clinic College of Medicine, Rochester, MN 55905, USA. Electronic address: tortorelli.silvia@mayo.edu.

Abstract

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.

KEYWORDS:

Dried blood spot; Lysophosphatidylcholine; Newborn screening; Peroxisomes; Tandem mass spectrometry; X-linked adrenoleukodystrophy

PMID:
25481105
DOI:
10.1016/j.ymgme.2014.11.013
[Indexed for MEDLINE]

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