*2.16. Amplex® Red Peroxidase Assay*

The rate of H2O2 reduction was assayed by the production of H2O2 in epimastigotes to H2O, which is stoichiometrically coupled (1:1) to the simultaneous oxidation of the non-fluorescent Amplex® Red probe to the fluorescent resorufin [48]. Briefly, 107 parasites/mL were incubated with 5 mM Tris-HCl (pH 7.4), 1.7 μM Amplex® Red (Invitrogen, Carlsbad, CA, USA), and 6.7 U/mL horseradish peroxidase (Sigma-Aldrich) in a final volume of 100 μL, for 30 min at room temperature. Fluorescence evolution was observed at excitation/emission wavelengths of 563/587 nm. H2O2 concentration was determined using a standard curve.
