Development of an Enzyme Immunoassay for Serum 16-Dehydropregnenolone
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概要
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We have developed an enzyme immunoassay(EIA)for serum 16-dehydropregnenolone(3β-hydroxy-5, 16-pregnadien-20-one ; 16-DHP). The antiserum against 16-DHP-3-hemisuccinate conjugated bovine serum albumin(16-DHP-3HS-BSA)was raised in rabbits. For use as an enzyme labeled antigen, 16-DHP-3HS was conjugated to alkaline phosphatase. The minimal amount of 16-DHP detected was 4 pg(0.013pmol)/assay and the measurable range was from 0.06-60ng/ml(0.191-191nmol/l). The intra-assay coefficient of variation(C.V.)was 4.1%(0.73±0.03ng/ml, mean±S.D., n=6), and inter-assay C.V.was 7.7%(0.13±0.01ng/ml, n=6). A liner relation was observed between the serum sample dilution and the 16-DHP concentration. For the recovery study, authentic 16-DHP was added to a serum sample(original concentration : 0.10-0.14ng/ml), and the recovery was found to be 94.4-96.8%(final 16-DHP concentrations calculated : 0.29-16.3ng/ml). To investigate the reliability of the present EIA, the values from our EIA were compared with those obtained by GC-MS. The 16-DHP concentration could not be measured in serum by GC-MS because of its sensitivity. Therefore, the conjugated steroid, 16-DHPS, was first enzymatically hydrolysed and then the 16-DHP measured by both methods. There was a good correlation between the levels determined by these methods(Pearson's correlation coefficient : r=0.927, p<0.001, y=0.74x+3.61, n=27). The serum concentrations of 16-DHP in neonates and umbilical vein were 0.53±0.09ng/ml and 0.88±0.61ng/ml, respectively. No 16-DHP was detected in serum from normal healthy adults using the present EIA. These results suggest that 16-DHP originates from the fetus and neonate.
- 公益社団法人日本薬学会の論文
- 2001-08-01
著者
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Saiki K
Central Analytical Laboratory
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Kobayashi Yoshiharu
Clinical Chemistry And Medicinal Chemistry Laboratories Kobe Women's College Of Pharmacy
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Kobayashi Yoshiharu
Clinical Chemistry Laboratory
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TAGAWA Noriko
Clinical Chemistry Laboratory
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SAIKI Kayoko
Central Analytical Laboratory
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