An enzymatic assay for adenosine 5′-triphosphate (ATP) and other nucleoside triphosphates and determination of the specific radioactivity of the terminal P

ATP can be assayed by transferring its terminal phosphate group to tritium-labeled glycerol with glycerokinase and measuring the radioactivity of the resultant glycerophosphate. The observed tritium activity is proportional to the ATP content of the sample. If the terminal P of ATP is radioactive, the 32P 3H ratio of the glycerophosphate is a selective measure of the specific activity of that P atom. The assay is useful over a range of 0.01–10 nmoles of ATP and is suitable for the assay of small volumes of tissue extracts. The assay is not specific for ATP and can be used to determine the other nucleoside triphosphates. However, the method can be modified with some loss of convenience to be specific for ATP.