DETECTION OF 100 MOLECULES OF PRODUCT FORMED IN A FUCOSYL-TRANSFERASEREACTION

We present the detection of 100 molecules of enzyme substrate and prod uct. A fucosyltransferase and a fucosidase were used to add and remove , respectively, a monosaccharide from a synthetic oligosaccharide fluo rescently labelled with tetramethylrhodamine. The reaction was followe d by use of capillary zone electrophoresis, to separate the product an d reactant, with laser-induced fluorescence detection. These are the m ost sensitive enzyme assays reported to date and six orders of magnitu de more sensitive than any reported for these two enzymes. This simple technology allows high-sensitivity determination of the activity of a ny enzyme for which a fluorescent substrate can be synthesized, and br ings within reach the ability to assay glycosyltransferase activities in single cells.