Kinetic determination of serum adenosine deaminase

A new kinetic method for the determination of serum adenosine deaminase (EC 3.5.4.4) is described, with adenosine as the substrate and nucleoside phosphorylase and xanthine oxidase as the reaction enzymes. Inosine is produced, which is converted to hypoxanthine. The hypoxanthine is oxidized to xanthine, which is further oxidized to uric acid. In these two reactions, blue 2,6-dichlorophenolindophenol is reduced to a colorless compound and the decrease in color is measured spectrophotometrically at 606 nm. The assay was automated by using a Cobas Mira analyzer. The automated assay had a CV of < 7%, and the calibration curve was linear from 10 to 120 U/L. The assay correlates well with an established method, based on detection of liberated NH3 with Berthelot's reaction. The reference interval (mean +/- 2 SD) was 14-34 U/L (mean 24 U/L, n = 84). The enzymatic method described is easily automated and seems to be suitable for the routine determination of adenosine deaminase in serum.