Rapid assay of N-acetyl-beta-D-glucosaminidase isoenzymes in urine by ion-exchange chromatography.
We describe a new method for separating and measuring urinary N-acetyl-beta-D-glucosaminidase isoenzymes by "high-performance" liquid chromatography. Isoenzymes are eluted from the anion-exchange column with a one-step linear gradient of NaCl solution. For continuous post-column quantification of the activities of these isoenzymes, we use an on-line post-column reactor and 4-methylumbelliferyl-N-acetyl-beta-D-glucosaminide as substrate; the methylumbelliferone formed in this reaction is quantified fluorimetrically. We discuss the effects of varying different components of the assay: NaCl concentration, the pH of the mobile phase and of the reaction reagent, substrate concentration, incubation temperature, and the geometry of the post-column reactor.
Nicot
G
G
Département de Pharmacologie Clinique, Centre Hospitalier Régional Universitaire, Hôpital Dupuytren, Limoges, France.
Lachâtre
G
G
Gonnet
C
C
Dupuy
J L
JL
Valette
J P
JP
D000118:Acetylglucosaminidase / Q000652:urine*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D005470:Fluorometry; D006596:Hexosaminidases / Q000652:urine*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000652:urine*; D016030:Kidney Transplantation
D007527:Isoenzymes; D006596:Hexosaminidases; D000118:Acetylglucosaminidase