Assessment of eleven adrenal steroids from a single serum sample by combination of automatic high-performance liquid chromatography and radioimmunoassay (HPLC-RIA)

Abstract

A radioimmunological method for the assessment of eleven adrenal steroids from one serum sample is presented. The method includes an extraction, purification and radioimmunological quantitation step. Automatic high-performance liquid chromatography (HPLC) is used as a Chromatographie techniquc for the purification step. The localization of steroid fractions during a Chromatographic run is triggered by the retention times established by chromatography of U.V.-detectable amounts of steroids prior to each batch. The main advantages of this automatic HPLC technique are (1) the high potency of resolution, (2) the high degree of reproducibility of retention times, (3) the gradient elution technique allowing “one run”-chromatography of compounds covering a great range of polarity, and (4) the automatic feature eliminating almost all manual operations. Specificity of the antisera raised against the steroids progesterone, pregnenolone, deoxycorticosterone, 17-OH-progesterone, 17-OH-pregnenolone, deoxycortisol, 18-OH-deoxycorticosterone, corticosterone, aldosterone, cortisol and 18-OH-corticoster-one were studied by cross-reactivity to known steroids and by monitoring the Chromatographic profiles of immunoreactivities arising in a normal serum sample. Base levels of serum steroids in normal man as well as their changes after adrenal stimulation and suppression were in accordance with the data reported in literature.