Abstract: The substrate solution consisting of 1 mM inosine in 50 mM phosphate buffer (pH 6.0) was pre-incubated at 37°C for ~5 min before addition of erythrocyte lysate (50 µL). Following incubation for 10 min, the reaction was stopped with 4 M HClO4. The resulting solution was mixed with 2 M K2CO3 (to remove KClO4) and centrifuged and a portion of the supernatant solution was merged (at 0.25 mL min-1) in the flow injection analysis sytem with 0.1 M Tris - HCl buffer (pH 8) as carrier solution. The resulting stream was mixed in the coil with reagent solution [as for the carrier solution but containing 5 mM 3-(4-hydroxyphenyl)propionic acid] and the hypoxanthine present in the stream was degraded to H2O2 on columns of immobilized xanthine oxidase, urate oxidase and horse-radish peroxidase in series. The fluorescence intensity of the final solution was measured at 405 nm (excitation at 305 nm). The limit of determination for hypoxanthine was 0.3 pmol in a 20 µL injection. The method permits the assay of purine-nucleoside phosphorylase in 10 nl of erythrocytes.