Figure 3. Resolubilisation of indigoidine in the inorganic solvent DMSO enables a linear standard curve to be generated.

(A) Replicate reaction mixes containing 50 mM Tris-Cl pH 8.5, 20 mM MgCl_2, 12 mM ATP, 3 μM holo-BpsA, 5 mM L-glutamine and ddH₂O to a total volume of 40 μL were incubated for 1 h at 25 °C. Addition of DMSO to final concentrations of 80% or higher were found to maximise the solubility of indigoidine present in an aqueous solution, enabling more accurate quantification via measurement of absorbance at 590 nm. (B) The A₅₉₀ of a fully solubilised 200 μL solution of indigoidine in 95% DMSO (■) was found to diminish only slightly over a 10 h period, consistent with indigoidine undergoing a gradual conversion into the colourless leuco isoform. Data are the mean values from two independent experiments, each comprising three technical replicates, and error bars indicate standard error of the mean.