The activity of intracellular chemical reactions of molecules is measured
by the use of fluorescently labeled substrate molecules that undergo a
change in electrophoretic mobility upon chemical reaction such as that
catalyzed by an enzyme. Specificity is achieved by using labeled
substrate molecules that can be acted upon only by specific enzymes. Thus
the activity of a specific enzyme or class of enzymes can be determined.
Measurements are made with the intracellular presence of such substrate
molecules, at some time of interest, typically after exposure of the cell
to a stimulus that activates a particular enzymatic pathway. To ensure
accuracy, measurements must be made in a timely manner so as to minimize
chemical reactions occurring subsequent to the time of interest. Fast
controllable laser lysis is used to obtain the contents of a single cell
into which reporter substrate molecules have been introduced. The cell
contents are then subjected to capillary electrophoresis and enzymatic
activity is determined by comparing amounts of substrate molecules to
amounts of enzymatically altered substrate molecules which are separated
by the electrophoresis and identified by the presence of a fluorescent
label.