Fig. 2

Response of roGFP1 and roGFP1-Se147 to oxidation and reduction. a Mean ± SEM of the 405/470 ratio of HEK293 cells transfected with roGFP1-Se147pSel (grey, n = 52) and roGFP1-pSel (black, n = 220) treated with 3, 30, and 300 µM H₂O₂ followed by 3 mM DTT. b and c Representative pseudocolor image of 405/470 ratio of HEK293 cells transfected with roGFP1-pSel (b) and roGFP1-Se147pSel (c) in control conditions and after treatment with 300 µM H₂O₂. d normalized mean ± SEM 405/470 response of HEK293 expressing roGFP1-pSel to 3 µM to 3 mM H₂O₂ (n = 62 to 263) with reduced recording frequency. *Denotes significant difference at 240 s in normalized ratio between different treatment groups (one way ANOVA, p < 0.01). e Normalized mean ± SEM 405/470 response of HEK293 expressing roGFP1-Se147pSel to 30 nM to 300 µM H₂O₂ (n = 103 to 346) with reduced recording frequency. *Denotes significant difference at 240 s in normalized ratio between different treatment groups (one way ANOVA, p < 0.01). f Concentration-normalized response relationship of H₂O₂ treatment in roGFP1-pSel and roGFP1-Se124pSel expressing HEK293 cells fitted using a non-linear regression. EC₅₀ for roGFP1-pSel and roGFP1-Se147pSel was 2.0 × 10⁻⁵ (R² of 0.94) and 9.8 × 10⁻⁷ (R² of 0.97), respectively. g and h normalized mean ± SEM 405/470 response of HEK293 expressing roGFP1 constructs to 10 μM antimycin A (red lines), 0.1% ethanol vehicle (green lines) and buffer (blue lines). g roGFP1-pSel (n = 221–363). h roGFP1-Se147pSel (n = 237–346). *Denotes significant difference in normalized ratio between different treatment groups (repeated measures ANOVA, p < 0.01). n.s. denotes no significant difference between groups (p > 0.05)