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Clinical Chemistry 35: 2322-2325, 1989;
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Clinical Chemistry, Vol 35, 2322-2325, Copyright © 1989 by American Association for Clinical Chemistry

Sensitive, simple staining method for use in electrophoretic determination of amylase isoenzymes in serum

F Mashige, N Hisae, S Toyota, S Yoshinoya and A Ohkubo
Department of Laboratory Medicine, Faculty of Medicine, University of Tokyo, Japan.

We have developed a sensitive and simple staining method for use in electrophoretic analysis of serum alpha-amylase (EC 3.2.1.1) isoenzymes. The principle of this method is as follows. alpha-Amylase hydrolyzes 4-nitrophenylmaltoheptaoside to generate oligosaccharide, which is then converted to gluconolactone in the presence of oligosaccharide dehydrogenase (no EC no. assigned), with concomitant reduction of 1-methoxy-5-methylphenazinium methylsulfate (1-m-PMS) to 1- m-PMSH. The hydrogen in 1-m-PMSH is then transferred to 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide to yield formazan. Each of the isoenzymes can then be measured densitometrically. The mean (and SD) values for total amylase, P1, S1, and S2 as determined by this method with sera from 25 healthy adults in fasting were 251 (64), 104 (35), 126 (40), and 22 (11) U/L, respectively. Between-assay CVs (n = 10) for determinations of P1, S1, and S2 were 3.54%, 4.03%, and 7.01%, respectively.




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Copyright © 1989 by the American Association for Clinical Chemistry.