Delayed luminescence analysis (DLA) of purine and pyrimidine ribose and deoxyribose nucleotide triphosphates in picomole quantities

A discovery is reported of a new system that enables one to quantitate the amounts of separated nucleotide triphosphates in picomole quantities. This system of delayed luminescence analysis (DLA) is sensitive to both purine and pyrimidine ribose and deoxyribose nucleotide triphosphates. A crude luciferin-luciferase (substrate-enzyme) preparation from firefly lanterns, in the presence of nucleotide triphosphate, is utilized to generate light that is detected by a liquid scintillation counter with the coincidence of the photomultiplier tubes turned off. Light is produced in a delayed fashion, the maximum emission being dependent on the type of nucleotide. Purine nucleotides (GTP, ITP, dATP, dGTP) give maximal light emission at approximately 2 mins; with the pyrimidine nucleotides the time required for maximal light emission was 5 min for UTP, dUTP, and TTP, 10 min for CTP, and 12 min for dCTP. A linear relationship on a log-log plot of light emission vs. concentration of nucleotide is demonstrated with ITP, dATP, UTP, and CTP.