A firefly luciferase assay for determination of cytidine 5'-triphosphate in biological samples.
A routine assay for CTP in cell and tissue extracts using crude firefly lantern preparations is described. ATP, GTP, and UTP are removed by incubation of the samples with a mixture of 3-phosphoglycerate kinase, hexokinase, glucose-6-phosphate dehydrogenase, and UDP-glucose pyrophosphorylase in the presence of ADP. The method is sensitive (greater than 30 nM CTP), inexpensive, and reproducible. No chromatographic purification of the biological samples or of the firefly extract is necessary. Corrections must be made for some loss of CTP during the enzymatic incubation and for the background luminescence of ADP. The applicability of the assay is tested with extracts from the cyanobacterium Anacystis nidulans.
D000244:Adenosine Diphosphate / Q000032:analysis; D000255:Adenosine Triphosphate / Q000302:isolation & purification; D000818:Animals; D001517:Coleoptera / Q000201:enzymology; D000458:Cyanobacteria / Q000032:analysis; D003570:Cytidine Triphosphate / Q000032:analysis*; D003597:Cytosine Nucleotides / Q000032:analysis*; D006160:Guanosine Triphosphate / Q000302:isolation & purification; D006868:Hydrolysis; D007700:Kinetics; D008156:Luciferases; D008163:Luminescent Measurements; D010783:Photometry; D014544:Uridine Triphosphate / Q000302:isolation & purification
D003597:Cytosine Nucleotides; D000244:Adenosine Diphosphate; D003570:Cytidine Triphosphate; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D008156:Luciferases; D014544:Uridine Triphosphate