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Purification of L‐Glutamate decarboxylase from rabbit brain and preparation of a monospecific antiserum
Author(s) -
Brandon C.
Publication year - 1986
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/jnr.490150308
Subject(s) - antiserum , glutamate decarboxylase , chromatography , size exclusion chromatography , immunoelectrophoresis , biochemistry , chemistry , microbiology and biotechnology , enzyme , polyclonal antibodies , biology , antibody , immunology
L‐Glutamate decarboxylase (GAD), the enzyme responsible for the biosynthesis of gamma‐aminobutyric acid, has been purified from rabbit brain and used for the production of a monospecific rabbit antiserum. The enzyme was purified approximately 500‐fold by a combination of ion exchange, hydrophobic, hydroxyapatite, and gel filtration column chromatography, and preparative polyacrylamide gel electrophoresis. The preparation thus obtained was injected into a rabbit, producing a polyspecific antiserum. This first antiserum was used for the preparation of an immunoprecipitate containing GAD; injections of the immunoprecipitate into a second rabbit led to the production of a second, monospecific anti‐GAD serum. The specificity of the antiserum for GAD has been demonstrated by enzyme precipitation, immunodiffusion, line immunoelectrophoresis, gel filtration chromatography of GAD/anti‐GAD immune complexes, and immunocytochemistry in the rat cerebellum.