Heme Oxygenase 1 (HO1) is a stress-inducible rate-limiting enzyme in heme catabolism, catalyzing oxidative cleavage of heme into biliverdin, ferrous iron and carbon monoxide. As a pivotal cytoprotective protein, it is highly upregulated under oxidative stress, hypoxia and inflammation, exerting robust anti-oxidative, anti-inflammatory and anti-apoptotic effects by scavenging reactive oxygen species and maintaining iron homeostasis. Widely expressed in the spleen, liver and vascular tissues, HO1 modulates macrophage polarization and tissue repair, and its dysregulation is linked to cardiovascular diseases, neurodegeneration and cancer, making it a key therapeutic target for oxidative stress-related pathologies. HO1 and BLVRB colocalize in iron-rich regions and interact to mediate heme degradation and redox balance in macrophages.Briefly, human BLVRB was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to HO1-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-BLVRB pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody for 1h at 37℃, wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µL stop solution to the wells and read at 450/630nm immediately. Measured by its binding ability in a functional ELISA. When Recombinant mouse HO1 is lmmobilized at 2 µg/mL(100 uLwell), the concentration of human BLVRB that produces 50% optimal bindingresponse is found to be approximately 0.019 µg/mL.