Work simplification of a competitive protein binding analysis for serum thyroxine a study of some analytical variables affecting accuracy and precision

Abstract

Some variables affecting the accuracy and precision of a previously described work simplified procedure for the competitive protein binding analysis of serum thyroxine have been investigated. Ethanol is preferred to ethanol-ammonia mixtures for the extraction of thyroxine from serum as it introduces the minimum interfering material into the assay. The use of various inhibitors of thyroxine binding to proteins other than thyroxine binding globulin has been investigated. None showed any advantage over the use of barbiturate alone. Conditions for obtaining maximum slope in the standard response curve are considered, with respect to buffer composition, concentration of binding protein, and time for equilibration and resin separation of protein bound and non protein bound thyroxine. Theoretical assessment of the errors in the analysis indicates that at low serum thyroxine concentrations, the error in interpolating results from the standard response curve constitutes a major part of the overall analytical error, while at medium or high concentrations, the error in the recovery step is more important.