SPÁČIL, Zdeněk, H TATIP2AKA, M BARCENAS, CR SCOTT, F TURECEK and MH GELB. High-Throughput Assay of 9 Lysosomal Enzymes for Newborn Screening. Clinical Chemistry. Washington, USA: Amer. Assoc. Clinical Chemistry, 2013, vol. 59, No 3, p. 502-511. ISSN 0009-9147. Available from: https://dx.doi.org/10.1373/clinchem.2012.189936.
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Original name High-Throughput Assay of 9 Lysosomal Enzymes for Newborn Screening
Edition Clinical Chemistry, Washington, USA, Amer. Assoc. Clinical Chemistry, 2013, 0009-9147.
Other information
Original language English
Type of outcome Article in a journal
Confidentiality degree is not subject to a state or trade secret
Impact factor Impact factor: 7.768
Doi http://dx.doi.org/10.1373/clinchem.2012.189936
UT WoS 000317337600010
Changed by Changed by: PharmDr. Zdeněk Spáčil, Ph.D., učo 238088. Changed: 29/9/2017 13:51.
BACKGROUND: There is interest in newborn screening of lysosomal storage diseases (LSDs) because of the availability of treatments. Pilot studies have used tandem mass spectrometry with flow injection of samples to achieve multiplex detection of enzyme products. We report a multiplexing method of 9 enzymatic assays that uses HPLC-tandem mass spectrometry (MS/MS). METHODS: The assay of 9 enzymes was carried out in 1 or 2 buffers with a cassette of substrates and internal standards and 1 or 2 punches of a dried blood spot (DBS) from a newborn screening card as the source of enzymes. The pre-HPLC-MS/MS sample preparation required only 4 liquid transfers before injection into a dual-column HPLC equipped with switching valves to direct the flow to separation and column equilibration. Product-specific and internal standard-specific ion fragmentations were used for MS/MS quantification in the selected reaction monitoring mode. RESULTS: Analysis of blood spots from 58 random newborns and lysosomal storage disease-affected patients showed that the assay readily distinguished affected from nonaffected individuals. The time per 9-plex analysis (1.8 min) was sufficiently short to be compatible with the workflow of newborn screening laboratories. CONCLUSIONS: HPLC-MS/MS provides a viable alternative to flow-injection MS/MS for the quantification of lysosomal enzyme activities. It is possible to assay 9 lysosomal enzymes using 1 or 2 reaction buffers, thus minimizing the number of separate incubations necessary. (C) 2012 American Association for Clinical Chemistry
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