Abstract
Recently the laser-nephelometer, which is especially designed for the measurement of the light scattered from immunoprecipitate in solution, has become available, and nephelometric immunoassay has become widely used for the determination of precipitating antigen such as serum protein. We expected that nephelometry would provide a method also for the determination of hapten (non-precipitating antigen). The assay principle is based on that hapten (monoantigenic small molecule) forms soluble immunocomplex which does not scatter the light whereas polyhaptenic molecule forms insoluble immunoprecipitate which scatters the light. Therefore, when the hapten competes for the binding site of the antibody with the haptenic moiety of the polyhaptenic molecule, the hapten can be determined by the measurement of the decrease of the scattered light due to the inhibition of the immunoprecipitation of the polyhaptenic molecule.
The sensitivity of this “competitive nephelometric immunoassay” method was not expected to be high. However therapeutic blood levels of some drugs were thought to be high enough for the sensitivity, and the monitoring of some drugs in patient body fluid is clinically useful. Therefore we used some drugs as model molecules to be determined, and used serum albumin as the hapten-carrier of the polyhaptenic molecule. The assay conditions of this method were studied, and we have developed an accurate, simple and rapid method for the determination of some drugs in human body fluid. The patients' plasma specimens were assayed, and the values correlated well to those determined by the high-performance liquid chromatography (correlation coefficient: 0.971 for theophylline, 0.983 for phenobarbital, 0.987 for diphenylhydantoin).
