Enzymatic assay in a lab-on-a-disk format to measure free piperacillin concentration in serum

This work describes a method based on an enzymatic assay to measure the free concentration of piperacillin (a [Formula: see text]-lactam antibiotics) in blood serum. The assay is based on the release of fluorescent umbelliferone, upon hydrolysis by BlaP99 [Formula: see text]-lactamase of a substrate in competition with piperacillin. The assay is implemented in a lab-on-a-disk (LoaD) setup, in which samples and reagents are manipulated with robust centrifugal microfluidic techniques. To encounter the needs and constraints of Therapeutic Drug Monitoring (TDM) at bedside, the system is entirely automated and miniaturized. Within a few minutes and from a few microliters of serum, it provides piperacillin concentration measurements in a clinically meaningful range. This paper describes the design and characterization of the chemical kinetics and the microfluidic strategy underlying this assay. The data are compared to measurements made in well-plates with a conventional method.