Figure 1: Characterization of reversible reaction-based glutathione probe RT.

(a) The reversible Michael addition reaction between RT and GSH. The function of each moiety in RT is highlighted (QY: quantum yield). (b) Ultraviolet–visible (unshaded) and fluorescence (shaded) spectra of RT (green) and RT-GSH (blue), the GSH adduct. (c) Linear relationship between F₄₀₅/F₄₈₈ and GSH concentrations. F₄₀₅ and F₄₈₈ are the fluorescence intensities for RT-GSH and RT, respectively. (d) Dynamic ranges of Grx1-roGFP2 and RT probe. Fluorescence ratio changes on treatment with different concentrations of H₂O₂ in HeLa cells is measured, R_60s/R_0s is calculated as the ratio R value 60 s after H₂O₂ treatment divided by the R value before treatment. Each point is the mean value of >8 cells analysed from 2 independent experiments. Error bars represent s.e.m.