A simple and rapid radiochemical choline acetyltransferase (ChAT) assay screening test.
A simple radiochemical choline acetyltransferase (ChAT) assay screening test was developed by measuring for [(3)H]acetylcholine ([(3)H]ACh) formed from 0.2 mM [(3)H]acetyl-coenzyme A ([(3)H]acetyl-CoA) and 1 mM choline by 0.2 mg of rat brain homogenates containing ChAT into 96-well microplates. A simple and rapid procedure for isolating [(3)H]ACh from the incubation mixture into 96-well microplates was achieved by using a sodium tetraphenylboron (Kalibor) solution (in ethyl acetate, 0.75%, w/v) and a hydrophobic liquid scintillator mixture (1:5, v/v, 0.2 mL) as an extraction solvent. The benefits of this radiochemical method using 96-well microplates are as follows: (1) this method is reliable and reproducible; (2) many samples can be examined at the same time by this method; (3) this method is economical and effective in reducing radioactive waste. The development of a new simple radiochemical ChAT assay screening test is the first stage of development of radiolabeled ChAT mapping agent.
Shiba
Kazuhiro
K
Division of Tracer Kinetics, Advanced Science Research Center, Kanazawa University, 13-1 Takara-machi, Kanazawa 920-8640, Ishikawa, Japan. shiba@med.kanazawa-u.ac.jp
Ogawa
Kazuma
K
Kinuya
Seigo
S
Yajima
Kazuyoshi
K
Mori
Hirofumi
H
D000109:Acetylcholine / Q000493:pharmacokinetics; D000818:Animals; D001921:Brain / Q000201:enzymology*; D001923:Brain Chemistry; D002795:Choline O-Acetyltransferase / Q000378:metabolism*; D011854:Radiochemistry / Q000379:methods*; D051381:Rats; D013997:Time Factors; D014018:Tissue Distribution; D014316:Tritium / Q000493:pharmacokinetics
D014316:Tritium; D002795:Choline O-Acetyltransferase; D000109:Acetylcholine