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Clinical Chemistry 20: 324-327, 1974;
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Clinical Chemistry, Vol 20, 324-327, Copyright © 1974 by the American Association for Clinical Chemistry

Chromogenic-Substrate Method for Demonstrating Multiple Forms of Alpha-Amylase after Electrophoresis

A. M. Spiekerman 1, P. Perry 1, N. C. Hightower 1, and F. F. Hall 1

1 Department of Research, Scott and White Memorial Hospital, Temple, Tex. 76501.

A chromogenic-substrate method is described for demonstrating multiple forms of alpha-amylase (isoamylases) after they have been electrophoretically separated in a continuous-buffer, slab-type, polyacrylamide-gel system. After electrophoresis, the various isoamylases are detected by layering the gel slabs onto thin-film plates of agar impregnated with Remazolbrilliant Blue starch. Isoamylases appear on the substrate plates as clear hydrolytic zones on a blue background of undigested substrate. After dehydration, the substrate plates can be stored indefinitely, and the polyacrylamide slabs are available for protein staining or elution of the isoamylase bands, if desired. This method has significant advantages: it is simple, substrate plates are easy to prepare, the plates can be permanently stored, optimum time required for isoamylase detection can be judged visually, and the method can be adapted for special two-dimensional electrophoretic studies.


Key Words: isoamylases • isoenzymes • Remazolbrilliant Blue starch • pancreatic and salivary amylases • electrophoresis on polyacrylamide gel, oneand two-dimensional

Accepted on December 3, 1973






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Copyright © 1974 by the American Association for Clinical Chemistry.