Clinical Chemistry, Vol 21, 715-718, Copyright © 1975 by the American Association for Clinical Chemistry

Fluorometric Assay of Serum -glutamyltransferase in Solution and on a Semi-Solid Surface

¹ Chemistry Department A, Building 207, Technical University of Denmark, 2800 Lyngby, Denmark.

We describe an enzymatic, fluorometric method for measuring the activity of serum -gIutamyItransferase (EC 2.3.2.2) in solution and on silicone rubber pads. N--L-Glutamyl--naphthylamide in tris(hydroxymethyl)aminomethane buffer is used as substrate. In solution, measurements are performed at 37 °C with use of a reaction volume of 3.0 ml, in Pyrex cuvets. Measurements on silicone rubber pads are also made at 37 °C, after establishing a stable substrate film by lyophilizing the reagent for the -glutamyltransferase assay on the surface of the pads. Only 10-50 µl of buffered substrate and serum containing the enzyme to make a total volume of 60 µl are necessary for each assay. The rate of appearance of -naphthylamine fluorescence (_ex = 344 nm; _em = 445 nm), liberated from N--L-glutamyl--naphthylamide by the enzymatic action of -glutamyltransferase, is measured and equated to its activity in serum. Calibration plots of the change in fluorescence per min vs. enzyme concentration for measurements in solution and on pads show a good proportionality in the range of 26.5-265 U/liter, and indicate the usefulness of these methods.