PAPS-reductase from Escherichia coli was employed to detect thioredoxins from pro- and eukaryotic organisms. A simple method for the isolation of this enzyme and properties of the enzymatic assay were described. A comparison between thioredoxins detected by the PAPS-reductase and the Fructose-bisphosphatase or NADP malate dehydrogenase was used to assess the validity of the test. The high cross-reactivity of the bacterial enzyme was useful in the purification of heterologous thioredoxins from spinach, Synechococcus, and Saccharomyces cerevisiae.