The synthesis of [γ-32P]ATP with high specific radioactivity was investigated by the enzymatic method. We utilized a part of the glycolytic pathway
which consists of three reaction steps. First, we examined the maximum radioactivity of H332PO4 without the decrease of labeling yield. Next, we examined the minimum amount of reagent without the decrease of labeling
yield, because some reagents contain the phosphorus species as impurity. In this paper, we propose the optimum condition for
synthesis of [γ-32P]ATP with high specific radioactivity. As our calculation, we will be able to obtain about 220 TBq/mmol specific radioactivity
using 1.5 GBq of H332PO4.