Preparation of highly concentrated super-hot [gamma-32P]ATP using small-scale ion-exchange chromatography.

In order to obtain a highly concentrated, pure and super-hot [gamma-32P]ATP, we improved the purification method of super-hot [gamma-32P]ATP which was synthesized by the method of Johnson and Walseth (1979). The super-hot [gamma-32P]ATP was synthesized in a relatively large volume (2 ml) of reaction mixture and purified using semi-micro scale anion exchange chromatography (Dowex 1 X 2, 60-70 microliters column volume). In combination with washing the reaction product with certain organic solvents, this chromatography technique makes it possible to obtain a highly concentrated and pure super-hot [gamma-32P]ATP (approx. 7,000 Ci/mmol; 20-30 mCi/ml) from [32P]Pi of any commercial source in a good yield.