RE: Enzyme Histochemistry for Glycogen Storage Diseases
Takeuchi (1958) Histochemical demonstration of branching enzyme
(amylo-1,4--1,6-transglucosidase) in animal tissues. J.Histochem.Cytochem.
50 mg. glucose-1-phosphate (Na or K
10 mg 5'-adenylic acid (free acid)
4 mg soluble glycogen
15 ml distilled water
10 ml 0.1 M pH 5,8 acetate buffer
1 g iodine
2 g potassium iodide
100 ml water
Tissue may be stored at -20
degrees C for up to 36 hr. but no longer.
1. Incubate 10-40 micron fresh frozen sections in the
incubating medium 1hr at room temperature.
2. Stain 5 min. in 10% Lugol's iodine
Sections can be mounted in glycerol gel
and examined immediately or mounted permanently. [A.A. Smith, E.M.
and H.Machida (1966) Durable
mounts of the iodine stain for the phosphorylase reaction. Stain Technol.
3. 0.1% iodine in 70% ethanol, 3 min.
4. 0.1% iodine in 95% ethanol, 5 min.
5. 0.1% iodine in absolute ethanol, 2 changes, 5 min.
6. 0.1% iodine in xylene, 2 changes, 5 min.
7. Mount in Clay-Adams "Histoclad" containing 1 mg iodine
per ml of resin. (Hartman-Leddon's "Histoclad"
can substituted. Neither "Permount" nor balsam works.)
Red-brown deposits indicate strong branching enzyme activity. Brown
deposits indicate moderate branching enzyme activity.
gray, or blue deposits indicate little or no branching enzyme
Allen A. Smith, Ph.D.
School of Graduate
Podiatric Medicine and Surgery
Shores, Florida 33161-6695
Does anyone out there in histo land know of a procedure for staining
"brancher" and "debrancher" enzymes....or are these strictly metabolic tests
that have to be performed on ground up flash frozen tissue?
Michigan State University
College of Human
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