Radiochemical microassay for aspartate aminotransferase activity in the nervous system

Journal Article · Fri Jul 01 00:00:00 EDT 1988 · Anal. Biochem.; (United States)

DOI:https://doi.org/10.1016/0003-2697(88)90416-2· OSTI ID:6583371

Garrison, D; Beattie, J; Namboodiri, M A

A radiochemical procedure for measuring aspartate aminotransferase activity in the nervous system is described. The method is based on the exchange of tritium atoms at positions 2 and 3 of L-2,3-(/sup 3/H)aspartate with water when this amino acid is transaminated in the presence of alpha-ketoglutarate to form oxaloacetate. The tritiated water is separated from the radiolabeled aspartate by passing the reaction mixture over a cation exchange column. Confirmation that the radioactivity in the product is associated with water was obtained by separating it by anion exchange HPLC and by evaporation. The product formation is linear with time up to 120 min and with tissue in the 0.05- to 10-micrograms range. The apparent Km for aspartate in the rat brain homogenate is found to be 0.83 mM and that for alpha-ketoglutarate to be 0.12 mM. Methods that further improve the sensitivity of the assay are also discussed.

• Research Organization: Georgetown Univ., Washington, DC (USA)
• OSTI ID: 6583371
• Journal Information: Anal. Biochem.; (United States), Vol. 172:1
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
AMINOTRANSFERASES
ENZYME ACTIVITY
ASPARTIC ACID
BRAIN
ION EXCHANGE
LIQUID COLUMN CHROMATOGRAPHY
RADIOASSAY
RATS
SENSITIVITY ANALYSIS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
WATER
AMINO ACIDS
ANIMALS
BODY
CARBOXYLIC ACIDS
CENTRAL NERVOUS SYSTEM
CHROMATOGRAPHY
ENZYMES
HYDROGEN COMPOUNDS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MAMMALS
NERVOUS SYSTEM
NITROGEN TRANSFERASES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANS
OXYGEN COMPOUNDS
RODENTS
SEPARATION PROCESSES
TRANSFERASES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques