Fiber optic immunochemical sensor for continuous, reversible measurement of phenytoin

This fiber optic sensor, based on a homogeneous fluorescence energy-transfer immunoassay, operates in a continuous, reversible manner to quantify the anticonvulsant drug phenytoin. B-Phycoerythrin-phenytoin and Texas Red-labeled antibody to phenytoin were sealed inside a short length of cellulose dialysis tubing, which was cemented to the distal end of an optical fiber. When the sensor was alternately placed into solutions with various concentrations of free phenytoin, the drug crossed the dialysis membrane and displaced a fraction of the B-phycoerythrin-phenytoin from the antibody. The resulting change in fluorescence signal was measured with a fiber optic fluorometer. A typical competitive-binding calibration curve was seen between 5 and 500 mumol of phenytoin per liter. Equilibrium response time ranged from 5 to 30 min for different sensors. Replicate equilibrium measurements with one sensor, alternated eight times between two solutions of phenytoin, gave a CV of 2.1% (n = 16) at 0 mumol/L and 2.4% (n = 13) at 100 mumol/L. A reversible immunochemical sensor can be made that has a response time suitable for continuous concentration measurements.