4.3. Bradford Assay

Protein assay dye reagent concentrate (BioRad Laboratories, Veenendaal, The Netherlands) was diluted 5 times with MilliQ water and filtered through a paper filter. This diluted stock solution was freshly prepared before use and kept in the dark at 4 °C. The albumin calibration line was determined in the standard range of 200–1000 µg mL⁻¹ protein concentration according to the supplier-provided protocol. A low-concentration assay was used in cases of lower protein concentrations (<25 µg mL⁻¹). Samples were prepared by mixing 980 µL diluted stock solution and 20 µL protein sample (for low-concentration assay: 800 µL diluted stock solution and 200 µL protein sample) followed by incubation for 5–10 min at room temperature. Absorption at 595 nm was measured and plotted against the protein concentration. Diluted ωTA enzyme samples were then prepared and measured in the same fashion in order to determine their concentration using the albumin calibration line.