ADP detection using an enzyme-coupled reaction

In a method for determining ATPase activity in a sample by measuring ADP to estimate the extent of reaction of ATP, where the method comprises the steps of:

• combining the sample, with reagents comprising ATP, pyruvate kinase, phosphoenolpyruvate, pyruvate oxidase and agents for determining the production of hydrogen peroxide from the reaction of pyruvate catalyzed by pyruvate oxidase, said agents producing a detectable product;
• and determining said detectable product;
• the improvement which comprises:
• prior to adding the ATP and said sample to said reagents, combining catalase or peroxidase and a non-interfering substrate with said reagents other than ATP and at least one of:
• 1) with said ATP, correcting components selected from the group consisting of (a) creatine phosphokinase and phosphocreatine and (b) pyruvate kinase and phosphoenolpyruvate, or
• (2) with said phosphoenolpyruvate, correcting components selected from the group consisting of (a) lactate dehydrogenase (LDH) and reduced nicotinamide adenine dinucleotide (NADH), (b) glutamic pyruvic transaminase and glutamic acid and (c) pyruvate oxidase and wherein the amounts employed of said correcting components are substantially non-interfering in said determining of ATPase activity.