OBJECTIVES: The aim of the present paper is a careful re-examination of an automated spectrophotometric procedure for glutathione transferase (GST) activity in human serum described previously and used in many laboratories. DESIGN AND METHODS: GST activity in human serum has been assayed spectrophotometrically under various experimental conditions. Recombinant human GSTs and specific inhibitors were also used to check the possible occurrence of artifacts. RESULTS: Basal level of the enzyme calculated using this method turns out to be much higher than that found using RIA and ELISA procedures. Relevant pH-dependent artifacts deeply affect this spectrophotometric assay. Notably, spectral changes previously interpreted as a measure of basal activity, are mainly due to an increase of the spontaneous reaction between the two substrates. CONCLUSION: GST activity in normal serum cannot be correctly determined with the spectrophotometric assay described previously because of the very low enzyme concentration and the pH-dependent artifacts.
OBJECTIVES: The aim of the present paper is a careful re-examination of an automated spectrophotometric procedure for glutathione transferase (GST) activity in human serum described previously and used in many laboratories. DESIGN AND METHODS: GST activity in human serum has been assayed spectrophotometrically under various experimental conditions. Recombinant humanGSTs and specific inhibitors were also used to check the possible occurrence of artifacts. RESULTS: Basal level of the enzyme calculated using this method turns out to be much higher than that found using RIA and ELISA procedures. Relevant pH-dependent artifacts deeply affect this spectrophotometric assay. Notably, spectral changes previously interpreted as a measure of basal activity, are mainly due to an increase of the spontaneous reaction between the two substrates. CONCLUSION: GST activity in normal serum cannot be correctly determined with the spectrophotometric assay described previously because of the very low enzyme concentration and the pH-dependent artifacts.