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 >0.99). Intra- and interassay CV were 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.
| Original language | English |
|---|---|
| Pages (from-to) | 1830-1838 |
| Number of pages | 9 |
| Journal | Clinical Chemistry |
| Volume | 49 |
| Issue number | 11 |
| DOIs | |
| Publication status | Published - Nov 2003 |
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ASJC Scopus subject areas
- Clinical Biochemistry
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Capillary Electrophoresis in Diagnosis and Monitoring of Adenosine Deaminase Deficiency. / Carlucci, Filippo; Tabucchi, Antonella; Aiuti, Alessandro; Rosi, Francesca; Floccari, Federica; Pagani, Roberto; Marinello, Enrico.
In: Clinical Chemistry, Vol. 49, No. 11, 11.2003, p. 1830-1838.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Capillary Electrophoresis in Diagnosis and Monitoring of Adenosine Deaminase Deficiency
AU - Carlucci, Filippo
AU - Tabucchi, Antonella
AU - Aiuti, Alessandro
AU - Rosi, Francesca
AU - Floccari, Federica
AU - Pagani, Roberto
AU - Marinello, Enrico
PY - 2003/11
Y1 - 2003/11
N2 - 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 >0.99). Intra- and interassay CV were 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.
AB - 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 >0.99). Intra- and interassay CV were 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.
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UR - http://www.scopus.com/inward/citedby.url?scp=0242267944&partnerID=8YFLogxK
U2 - 10.1373/clinchem.2003.021576
DO - 10.1373/clinchem.2003.021576
M3 - Article
C2 - 14578314
AN - SCOPUS:0242267944
VL - 49
SP - 1830
EP - 1838
JO - Clinical Chemistry
JF - Clinical Chemistry
SN - 0009-9147
IS - 11
ER -