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Purification and immunochemical properties of choline acetyltransferase from human brain

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Abstract

Choline acetyltransferase (CAT) was purified to homogeneity from 363 g of human neostriatum by means of ammonium sulfate and protamine sulfate fractionation, followed by chromatography on DEAE-Sephadex, hydroxyapatite, phosphocellulose, and agarose-hexane-Co A columns. The final product migrated as a single component on 7.5% gels with or without SDS. It had a molecular weight of 66,000 daltons and a specific activity of 7.3 μmol acetylcholine formed per milligram protein per minute. Antibodies prepared in rabbits gave single precipitin lines against this protein on Ouchterlony immunodiffusion and immunoelectrophoresis plates. The CAT-anti-CAT IgG complex migrated as a single band on gel electrophoresis, establishing the monospecificity of the antibodies. Strong cross-reactivity to the IgG was obtained with CAT from rat, rabbit, and guinea pig, but only weak reactivity with chicken. Fab fragments were prepared from the rabbit IgG and were used to stain CAT-containing neurons in the spinal cord and nerve endings at the neuromuscular junction using the PAP technique.

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Authors and Affiliations

  1. Kinsmen Laboratory of Neurological Research Department of Psychiatry, University of British Columbia, V6T 1W5, British Columbia, Vancouver, Canada

    J. H. Peng, P. L. McGeer, H. Kimura, S. C. Sung & E. G. McGeer

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  1. J. H. Peng
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  2. P. L. McGeer
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  3. H. Kimura
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  4. S. C. Sung
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  5. E. G. McGeer
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Peng, J.H., McGeer, P.L., Kimura, H. et al. Purification and immunochemical properties of choline acetyltransferase from human brain. Neurochem Res 5, 943–962 (1980). https://doi.org/10.1007/BF00966135

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