Galectin 3 (GAL3) is a member of the lectin family, of which 14 mammalian galectins have been identified. It is also a member of the beta-galactoside-binding protein family that plays an important role in cell-cell adhesion, cell-matrix interactions, macrophage activation, angiogenesis, metastasis, apoptosis. The protein also has been demonstrated to be involved in cancer, inflammation and fibrosis, heart disease, and stroke. GAL3 is expressed in the nucleus, cytoplasm, mitochondrion, cell surface, and extracellular space. It also can agglutinate red blood. In this case, we choose rabbit erythrocyte (RaE) to assay its ability of agglutination. A general procedure for hemagglutination assay (HA) is as follows, two-fold dilute the recombinant rat GAL3 with 0.01M PBS (pH7.4), add 50μL a serial dilution of GAL3 to each well of a U or V-bottom shaped 96-well microtiter plate. The final well serves as a negative control with no GAL3, replace with 50μL 0.01M PBS. Then add 50μL 1% RaE to each well and mixed gently. The plate is incubated for 1-2 hours at room temperature. The results are shown in Figure 1. The minimal effective concentration of GAL3 is 2.5μg/mL. (A) 1% RaE treated with 2.5μg/mL GAL3 for 2h; (B) Negative control without GAL3.
Figure. The hemagglutination activity of recombinant rat GAL3.