RadioenzymaticMicroassayfor SimultaneousEstimationsof Dopamine, Norepinephrine,and Epinephrinein Plasma,Urine,and Tissues

With this sensitiveand specific radioenzymatiC assay pico-gram quantities of norepinephrine,epinephrine, and dopa-mine can be simultaneously determined directly in plasma, urine, and tissue. We discuss in detail the steps that are critical for success of the assay, as well as the rationale behind the use of specific reagents. The assay is appropriate for use in any routine clinical chemistry laboratory. This assay is sensitive to approximately 1 pg of catecholamine. As compared with a commercially available method, this assay issignificantlymoreeconomical and less time consuming. Addftlonal Keyphrases: catecholamines economics of labora-toty operation assay optimization Several radioenzymatic assays have been developed for measuring dopamine (DA), norepinephrine (NE), and epi-nephrine (E) in biological samples (1-12) based on the conversion of DA, NE, and E to their O-methylated tritiated derivatives by catechol-O-methyltransferase (COMT; EC 2.1.1.6) and radioactive S-adenosylmethionine (SAM).2 Al-though these radioenzymatic assays can accurately measure catecholamines in the picogram range, some difficulties still remain (13), primarily related to the low concentrations of catecholamine and to the presence of inhibitory substances in biological samples and in the partly purified COMT. By interfering with the enzymatic methylation reaction, these inhibitors cause a high variability in the recoveries of the end products. Here we describe certain modifications in the incubation, extraction, and purification procedures that decrease intra- and interassay variability; increase the sensitivity and specificity of the assay; and permit the simultaneous estimation of DA, NE, and E in less than 50 1iL of plasma, urine, or extract of tissue