Additional file 3: Figure S3. of TriPer, an optical probe tuned to the endoplasmic reticulum tracks changes in luminal H2O2

In vitro, H2O2-driven formation of disulfide-bonded high molecular weight TriPer species with divergent optical properties. (A) Coomassie-stained non-reducing and reducing SDS-PAGE of wild-type TriPer or its mutant variant lacking the peroxidatic cysteine (TriPer C199S ) following exposure to 1.5 μM of H2O2 for the indicated time period. Black arrow denotes disulfide-bonded high molecular weight TriPer species. (B) As in (A), but following a 15-min exposure to increasing concentrations of oxidized PDI (0–8 mM). Note the lack of the high molecular weight species in this sample and their prominence in the H2O2-treated sample, (A) above. (C) Excitation spectra (measured at emission 535 nm) of HyPer (orange trace) and TriPer (blue trace) for the different states of the probes, corresponding to phases 1–4 in Fig. 3c. (PDF 205 kb)