TY - JOUR
T1 - Direct non-radioactive assay of galactose-1-phosphate:uridyltransferase activity using high performance liquid chromatography
AU - Lindhout, Martijn
AU - Rubio-Gozalbo, M Estela
AU - Bakker, Jaap A.
AU - Bierau, Jorgen
PY - 2010/7/4
Y1 - 2010/7/4
N2 - Background Galactose-1-phosphate uridyltransferase (GALT) catalyses the conversion of galactose-1-phosphate (Gal-1-P) and UDP-glucose (UDP-Glc) into glucose-6-phosphate and UDP-galactose (UDP-Gal) Complete, or near complete, deficiency of GALT causes classic galactosaemia. The diagnosis is confirmed by measuring GALT activity in erythrocytes The most commonly used assays require radio labelled substrates or indirect coupled assays Methods: GALT activity was measured in erythrocyte lysates using optimal concentrations of the substrates galactose-1-phosphate and UDP-Glc UDP-Gal and UDP-Glc were separated using reversed-phase high performance liquid chromatography with UV detection Clinical validity was assessed using blood samples from galactosaemic patients Results UDP-Gal and UDP-Glc were separated with HPLC. The assay was linear with incubation times up 80 min and between 0 and 425 nmol haemoglobin Within-day and between-day imprecision at 50, 75 and 100% enzyme activity was
AB - Background Galactose-1-phosphate uridyltransferase (GALT) catalyses the conversion of galactose-1-phosphate (Gal-1-P) and UDP-glucose (UDP-Glc) into glucose-6-phosphate and UDP-galactose (UDP-Gal) Complete, or near complete, deficiency of GALT causes classic galactosaemia. The diagnosis is confirmed by measuring GALT activity in erythrocytes The most commonly used assays require radio labelled substrates or indirect coupled assays Methods: GALT activity was measured in erythrocyte lysates using optimal concentrations of the substrates galactose-1-phosphate and UDP-Glc UDP-Gal and UDP-Glc were separated using reversed-phase high performance liquid chromatography with UV detection Clinical validity was assessed using blood samples from galactosaemic patients Results UDP-Gal and UDP-Glc were separated with HPLC. The assay was linear with incubation times up 80 min and between 0 and 425 nmol haemoglobin Within-day and between-day imprecision at 50, 75 and 100% enzyme activity was
KW - Galactosaemia
KW - Galactose-1-phosphate uridyltransferase
U2 - 10.1016/j.cca.2010.03.032
DO - 10.1016/j.cca.2010.03.032
M3 - Article
C2 - 20359473
SN - 0009-8981
VL - 411
SP - 980
EP - 983
JO - Clinica Chimica Acta
JF - Clinica Chimica Acta
IS - 13-14
ER -