2.8. Myeloperoxidase Activity (MPO) Assay

The myeloperoxidase activity is used as a biochemical marker of polymorphonuclear leukocyte influx (mainly neutrophil) to the injured tissue. MPO activity was determined using an assay described previously [11, 12]. After six hours of application of croton oil, the MPO enzyme activity was assessed in the ear samples. Tissue samples were homogenized with a motor-driven homogenizer in acetate buffer (8 mM, pH 5.4) containing HTAB. For evaluation of MPO activity, the supernatant was incubated with TMB (18.4 mM) at 37°C for 3 min. The enzyme activity value was assessed colorimetrically at 630 nm using a microplate reader (Fisher Biotech BT, 2000). The results were expressed as optical density (OD)/mL of the sample.