MPO Activity Assay

The infiltration of neutrophil into the colon tissue was determined by MPO activity, as previously described.39 The colon tissues were homogenized and centrifuged. Then, the pellet was resuspended and centrifuged. The supernatant was harvested and added to the substrate containing o-dianisidine hydrochloride (0.167 mg/mL), 0.0005% hydrogen peroxide, and KH₂PO₄ (50 mM). The absorbance was measured at 460 nm immediately and recorded as the optical density (OD) value. The second OD value was read after 5 min. The changes of OD value in 5 min were represented as MPO activity, which were normalized to total protein concentration.