Human brain-type glycogen phosphorylase: quantitative localization in human tissues determined with an immunoassay system.
Glycogen phosphorylase (EC 2.4.1.1) from human brain tissue was purified to homogeneity. Antisera were developed in rabbits with purified phosphorylase as the immunogen. Antibodies were first affinity-purified with a column of brain phosphorylase-coupled Sepharose, and then the antibody fraction was adsorbed with a column of muscle phosphorylase-coupled Sepharose to remove antibodies reactive also with muscle phosphorylase. By using the specific antibodies, a sandwich-type immunoassay system for measurement of brain phosphorylase was prepared. The assay system consisted of polystyrene balls with immobilized antibrain phosphorylase F(ab')2 fragments and the same antibody Fab' fragments labeled with beta-D-galactosidase from Escherichia coli. The assay was sensitive and specific to the brain phosphorylase. The minimum detection limit of the assay was 0.1 ng/assay tube, and the cross-reactivity of the assay with muscle phosphorylase was less than 1%. Tissue concentrations of immunoreactive brain-type phosphorylase were estimated. The phosphorylase was present in the heart at as high a level as in the brain. The immunoreactivity for brain phosphorylase was distributed widely at a significant concentration in various peripheral tissues, such as the digestive tract, bladder, aorta, liver, and testis. Immunohistochemical localization of brain phosphorylase in the CNS revealed that the enzyme is present in most astrocytes and amyloid bodies, as well as in some neurons in the cerebral cortex and Golgi cells in the cerebellar cortex.
Kato
K
K
Department of Biochemistry, Institute for Developmental Research, Aichi, Japan.
Shimizu
A
A
Kurobe
N
N
Takashi
M
M
Koshikawa
T
T
D001921:Brain / Q000201:enzymology*; D006801:Humans; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000032:analysis*; D006005:Phosphorylases / Q000032:analysis*; D011786:Quality Control; D012016:Reference Values; D014018:Tissue Distribution
D007527:Isoenzymes; D006005:Phosphorylases