ABSTRACT
l-Glutamate decarboxylase (GAD) which catalyzes the conversion of l-glutamate to γ-aminobutyric acid, an important inhibitory neurotransmitter, is a specific marker for γ-aminobutyric agric neurons and their processes. The purification procedures involved the initial extraction of GAD activity from the whole brain or from the crude mitochondrial fraction, followed by ammonium sulfate fractionation or concentration by ultra filtration and a series of column chromatographies including diethylaminoethyl -cellulose, hydroxylapatite, and gel filtration and finally by preparative nondenaturing polyacrylamide gel electrophoresis. Once the purity of GAD preparations was established from extensive analyses, the purified GAD preparations were used as an antigen for the production of specific antibodies. In addition, the immunocytochemical staining of GAD using polyclonal anti-GAD and monoclonal anti-GAD will also be compared. The species-specificity of GAD was examined by the double diffusion, enzyme inhibition, and microcomplement fixation tests employing both antimouse brain GAD serum and an-ticatfish brain GAD serum.
