Abstract
The clinical value of high sensitivity assay methods for determining the levels of therapeutic drugs in biological fluids is now widely accepted both for monitoring treatment and for accurate pharmacokinetic studies. Radioimmunoassay has recently been applied to this area of clinical biochemistry. This thesis describes the development of radioimmunoassays for phenytoin and etorphine and discusses problems encountered in the developmental work of an assay for sulthiame. It also examines some of the critical factors in antibody and label production in general. A sensitive and specific radioimmunoassay for phenytoin has been developed and can be used to determine the amount of drug in micro-samples of blood. Sufficient serum for this assay can be collected from a finger-prick, an important factor when the patient is an infant epileptic or if vene-puncture is difficult. The radioimmunoassay has been applied to the study of the disappearance of phenytoin-from the blood of volunteers following a single oral dose, and has shown pharmacokinetic phenomena that have been undetected by other workers using less sensitive methods. Although the development of a radioimmunoassay for sulthiame was unsuccessful, it has exemplified many of the problems encountered in the raising of antibodies to, and the radioactive labelling of, small molecules. Using a morphine radioimmunoassay as a model, factors affecting the immune response to small molecules have been examined, as well as methods for increasing the immunogenicity of the drug-carrier conjugates and for influencing the specificity of the resulting antibodies. These principles have been applied to the development of a radioimmunoassay for etorphine to produce the first method for measuring the low levels of this drug present in biological fluids. A high specific activity radioactive label is essential for the development of a sensitive assay. For many drugs this can only be achieved effectively by using a [3]H-label; radioiodine, the other isotope frequently used in radioimmunoassays has not proved suitable for labelling these small compounds.