Materials and Methods

After extraction of the sample (onfilterpaper) in the counting vial, cortisol is assayed by adding 50 L of horse serum containing tritiated cortisol, and 2 mL of toluene scintillator, shaking for 20 mm, and counting the radioactivity in a liquid-scintillation counter at ambient temperature. The method can be applied to saliva, serum, or urine-directly as wet samples or dried on the filterpaper. Only inexpensivereagents,which are stable for monthsto years, are needed. Additional Keyphrases: steroidhormones assaysof samples dried on filter paper competitive-bindingassays scintillation fluidused to separate boundand free analyte urine saliva pediatricchemistry We have previously described some radiotransinassays for cortisol (1, 2) based on the principle of competitive binding to indigenous plasma proteins or “transins ” (e.g., transcortin, sex-hormone-binding globulin, thyroxin-bind-ing globulin). They differ chiefly with respect to procedures for deproteinizing the sample to be assayed (if it contains any binding-proteins) or for separating the bound and free moieties of the tracer in the final assay mixture. In the present method the samples, applied to small pieces of ifiter paper, are placed in counting vials and covered with methylene chloride to extract the cortisol. The paper, which retains the aqueous phase, is removed, the solvent is evapo-rated, and horse serum containing cortisol tracer is added, followed by scintillator. Cortisol extracted from the sample competes with tracer for binding sites on the transcortin of the horse plasma, the bound portion remaining in the aqueous phase while the displaced tracer passes into the scintillator phase, where its radioactivity is counted. The entire procedure is carried out at room temperature