Radioisotopic assay of femtomole quantities of total adenine nucleotides, ATP plus ADP, and AMP.
AMP is converted to ATP by incubating overnight with pyruvate kinase, phosphoenolpyruvate and adenylate kinase in the presence of endogenous ATP (ADP) as primer. In a subsequent incubation in the presence of pyruvate kinase, phosphoenolpyruvate, radioactive glucose and hexokinase, ATP and ADP are estimated together by coupling their recycling to the formation of glucose 6-phosphate. The latter is separated by precipitation using 76% (v/v) acetone for radioactivity measurement in the same Eppendorf tube. The sensitivity of these simple procedures matches or exceeds those of luciferase methods of nucleotide determination.
D000227:Adenine Nucleotides / Q000032:analysis*; D000244:Adenosine Diphosphate / Q000032:analysis; D000249:Adenosine Monophosphate / Q000032:analysis; D000255:Adenosine Triphosphate / Q000032:analysis; D000818:Animals; D002250:Carbon Radioisotopes; D066298:In Vitro Techniques; D008832:Microchemistry / Q000379:methods; D008930:Mitochondria, Liver / Q000032:analysis; D051381:Rats
D000227:Adenine Nucleotides; D002250:Carbon Radioisotopes; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate