Chromatographyを用いない血漿Aldosteroneの測定法とその検討
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Recently, non-chromatographic methods for the radioimmunoassay of plasma aldosterone have been reported. In this paper, we describe a new modified method for the radioimmunoassay of plasma aldosterone using Fuller's earth, charcoal and anti-aldosterone serum to purify aldosterone in plasma.<BR><SUP>3</SUP>H-aldosterone was added to 2 ml of plasma samples. Then, the free steroid hormone in the plasma was absorbed on 50 mg of the Fuller's earth and extracted with dichloromethane. The dichloromethane was dried. For the purification of aldosterone, AbI (1 : 10,000 diluted antibody to aldosterone) was added to each of the tubes and incubated for 2 hours at room temperature. Separation of free from bound fractions was performed by dextran-coated charcoal. The supernatant containing the bound fraction was transferred into small test tubes and extracted with dichloromethane. The extract was used for calculating recovery and for assay. A standard curve was constructed by adding <SUP>3</SUP>H-aldosterone and various amounts of standard aldosterone. Abll (1 : 200,000 diluted antibody to aldosterone) was added to standard tubes and sample tubes, and these tubes were incubated. The free fraction was separated by the addition of saturated ammonium sulfate and counted.<BR>In this method, the mean recovery of <SUP>3</SUP>H-aldosterone for 70 specimens was 28.6±7.98 % (S.D.). While, the recovery of 3H-hydrocortisone for 10 specimens was 0.03±0.03 % (S.D.). The water blank was assayed 0 ng/dl (n=4). The sensitivity of the standard curve was 2 pg. Accuracy, intra-assay precision, and specificity were examined by comparing the results with a method using our radioimmunoassay containing a paper chromatographic purification step, and determining the aldosterone content of plasma from various patients and plasma to which had been added different amounts of standard aldosterone to pre-extracted plasma. The equation for the analysis of variance on 20 specimens was Y=0.72X+1.874, r=0.925. The discrepancy between the two methods was not significant. In 9 normal subjects, the plasma aldosterone content ranged from 0 to 9.6 ng/dl. In 6 patients with primary aldosteronism, it ranged from 19.2 ng/dl to 50.3 ng/dl. Also, in 4 patients with primary aldosteronism and in 4 normal subjects, circadian rhythms of plasma aldosterone showed a higher content in the morning than in the evening.<BR>This method eliminated chromatography and easily enabled one technician to process a great number of samples in 5-6 hours.
- 日本内分泌学会の論文
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