Measurement of Free Phenytoin in Blood with a Self-Contained Fiber-Optic Immunosensor

We describe the measurement of the anticonvulsant drug phenytoin (5,5-diphenylhydantoin) in human blood and plasma using a self-contained fiber-optic immunosensor which gave a reversible response to changes in concentration. No regenerative treatment of the immunosensor was required between measurements. The analytical signal depended on the degree of energy transfer from B-phycoerythrin labeled with phenytoin to Texas Red labeled anti-phenytoin antibody. Dextran 70K was added to the reagent system to equalize the oncotic pressure across the encapsulation membrane. A gas chromatography reference method was used to measure free drug concentrations. Regression analysis for plasma samples gave the relationship (sensor) = 1.02 (reference) + 0.07 microM; Syx = 1.00 microM; r = 0.953; sensor mean 4.45 microM; reference mean 4.31 microM. Sensor performance in plasma and whole blood was essentially equivalent. We demonstrate the feasibility of measuring free phenytoin directly in blood and suggest that the sensor design is generally applicable for the measurement of other haptens in blood.