In:
Clinical Chemistry, Oxford University Press (OUP), Vol. 49, No. 11 ( 2003-11-01), p. 1830-1838
Abstract:
Background: The diagnosis and monitoring of severe combined immunodeficiency disease (SCID) attributable to adenosine deaminase (ADA) deficiency requires measurements of ADA, purine nucleoside phosphorylase (PNP), and S-adenosyl-l-homocysteine-hydrolase (SAHH) activity and of deoxyadenosine metabolites. We developed capillary electrophoresis (CE) methods for the detection of key diagnostic metabolites and evaluation of enzyme activities. Methods: Deoxyadenosine metabolites were separated in 30 mmol/L sodium borate–10 mmol/L sodium dodecyl sulfate (pH 9.80) at 25 °C on a 60-cm uncoated capillary. For determination of enzyme activities, substrate–product separation and measurements were carried out in 20 mmol/L sodium borate (pH 10.00) at 25 °C on a 42-cm uncoated capillary. Results: Deoxynucleotides and deoxyadenosine were readily detectable in erythrocytes and urine, respectively. Both methods were linear in the range 2–500 μmol/L (r & gt;0.99). Intra- and interassay CV were & lt;4%. Enzyme activities were linear with respect to sample amounts in the incubation mixture and to incubation time (r & gt;0.99 for both). In erythrocytes from healthy individuals, mean (SD) ADA activity was 5619 (2584) nmol/s per liter of packed cells. In erythrocytes of SCID patients at diagnosis, ADA activity was 56.9 (48.3) nmol/s per liter of packed cells; SAHH activity was also much reduced. PNP activity was similar in patients and controls. Conclusions: CE can be used to test ADA deficiency and enables rapid assessment of ADA expression in hematopoietic cells of SCID patients during therapy.
Type of Medium:
Online Resource
ISSN:
0009-9147
,
1530-8561
DOI:
10.1373/clinchem.2003.021576
Language:
English
Publisher:
Oxford University Press (OUP)
Publication Date:
2003
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