In the first step, samples are incubated in the antibody-coated microtiter plate. During the second step, the plate is washed and incubated with the POD-labeled HO-1 antibody. The reaction between POD and the substrate (H2O2, TMBZ) results in a color development. The amount of sample HO-1 is determined by measuring absorbance using an EIA plate reader. Accurate HO-1 sample concentrations can be determined by comparing their specific absorbances with the absorbance obtained for the Standard plotted on a standard curve.
Heme oxygenase is a microsomal enzyme which functions to degrade heme, a prosthetic group of heme proteins (e.g. hemoglobin), into the bile pigment biliverdin (which is converted to bilirubin), carbon monoxide, and reduced iron (Fe2+). Bilirubin has an anti-inflammatory effect in the body through its strong radical scavenging activity, whereas carbon monoxide has a vasodilative effect on organ blood flow.
Heme oxygenase exists as at least two isozymes (heme oxygenase-1 and heme oxygenase-2), which differ in their tissue expression and inducibility. Heme oxygenase-1 (HO-1) is an enzyme whose expression is induced intracellularly in response to various types of stress, and monitoring of HO-1 may prove useful for identifying stress induction.
