FIG. 1.

[H₂O₂] calibration with different electrochemical techniques. Experiments were conducted in PBS-I buffer, using a 10 μm Pt-UME versus Ag/AgCl. (A) CVs with 0 (black) and 20 μM H₂O₂ (red). Potential window from -0.1 to +1.0 V; scan rate 50 mV/s. In the inset (potential window of 0.6–0.7 V at a larger scale), the lower detection limit for [H₂O₂] of 50 nM (red trace; black trace 0 H₂O₂) at a scan rate of 20 mV/s is exemplified. Green arrows indicate forward and backward directions of cycling. (B) LSVs with 0 (black) and 35 μM H₂O₂ (red). Forward scanning was conducted from -0.1 to +1.0 V at a scan rate of 50 mV/s. Then, potential was reduced in one quick step to -0.1 V, clamped for 3 s, before forward scanning started again. (C) Representative CA measurement at 650 mV under pulsed additions of H₂O₂ (5 μM steps; arrows) to cell-free buffer (example for n = 10 similar experiments). (D) Current–dose relationship for H₂O₂ measured in CV at different potential windows described in the box (scan rate 50 mV/s). ΔI values represent current values sampled at 650 mV subtracted by respective basal current values of the CV before addition of H₂O₂. For each experiment with different potential window and/or different [H₂O₂] current values (ΔI₆₅₀) of sweep numbers 2–10 were averaged. Mean ± SEM (n = 5) is shown. (E) Current–dose relationship for H₂O₂ measured in LSV at different potential windows described in the box [scan rate 50 mV/s; protocol as in (B)]. Current values (ΔI₆₅₀) were analyzed as for CVs in (D). Mean ± SEM (n = 4) is shown. (F) Current–dose relationship measure in CA for [H₂O₂] from 500 nM to 1 mM (cell free, n = 10; red inset n = 3 for [H₂O₂] of 0–5 μM). For calculations, current values (I₆₅₀) of the respective approximate linear steady-state phase after each H₂O₂ addition were averaged. Mean ± SEM is shown. CA, chronoamperometric; CV, cyclic voltammogram or cyclic voltammetry; [H₂O₂], local hydrogen peroxide concentration; LSV, linear scan voltammogram; UME, ultramicroelectrode.