Two staining methods for selectively detecting isomaltase and maltase activity in electrophoresis gels.
Two methods for specifically detecting maltase, alpha-glucosidase, or isomaltase activity in electrophoresis gels are described. Both systems couple the formation of glucose by enzyme action on maltose or isomaltose to the generation of a colored product. System A uses an agarose overlay which contains substrate, glucose oxidase, peroxidase, 2,4-dichlorophenol, and 4-L-amino-phenazone. A purple color is produced at the site of enzyme activity. No hazardous chemicals are used at any stage. The stain is simple, rapid, sensitive, and inexpensive and does not interfere with subsequent protein staining. However, the stain is not permanent. System B was developed to give a permanent stain. The gel is overlaid with agarose containing substrate, glucose oxidase, phenazine methosulfate, and nitroblue tetrazolium. Glucose production results in the nitroblue tetrazolium being oxidized to an insoluble formazan with a dark blue color. This stain is also sensitive, rapid, and inexpensive but does use hazardous chemicals and if overstaining occurs this can interfere with subsequent protein staining. Neither system inactivates the localized enzymes which can be recovered from the gel if desired.
Finlayson
S D
SD
Department of Bioscience and Biotechnology, University of Strathclyde, Glasgow, Great Britain.
Moore
P A
PA
Johnston
J R
JR
Berry
D R
DR
D000632:Aminopyrine; D004586:Electrophoresis; D004587:Electrophoresis, Agar Gel; D005562:Formazans / Q000032:analysis; D005947:Glucose / Q000096:biosynthesis; D006026:Glycoside Hydrolases / Q000032:analysis*; D007534:Isomaltose / Q000378:metabolism; D008320:Maltose / Q000378:metabolism; D009580:Nitroblue Tetrazolium; D009834:Oligo-1,6-Glucosidase / Q000032:analysis*; D010084:Oxidation-Reduction; D013194:Staining and Labeling; D013778:Tetrazolium Salts; D000520:alpha-Glucosidases / Q000032:analysis*
D005562:Formazans; D013778:Tetrazolium Salts; D000632:Aminopyrine; D009580:Nitroblue Tetrazolium; D007534:Isomaltose; D008320:Maltose; D006026:Glycoside Hydrolases; D009834:Oligo-1,6-Glucosidase; D000520:alpha-Glucosidases; D005947:Glucose