Galectin-12 is a member of a family of mammalian lectins known as galectins. The galectins constitute a large family of carbohydrate-binding proteins that function in many systems both intracellularly and following secretion. Galectins contain either one or two carbohydrate recognition domains (CRR) which mediate recognition of N-acetyl-lactosamine-containing glycoproteins. Individual galectins differ in their tissue distribution and in their carbohydrate-binding specificities. Galectin-12 is predominantly expressed in adipose tissue and detected also in macrophages and other leukocytes. It plays an important role in cell-cell adhesion, cell-matrix interactions, macrophage activation, angiogenesis, metastasis, apoptosis. In this case, we chose rabbit erythrocyte (RaE) to assay its ability of agglutination. A general procedure for hemagglutination assay (or haemagglutination assay; HA) is as follows, two-fold dilute the recombinant rat GAL12 with 0.9% sodium chloride injection, add 50μL a serial dilution of GAL12 to each well of a U or V-bottom shaped 96-well microtiter plate. The final well serves as a negative control with no GAL12, replace with 50μL 0.9% sodium chloride injection. Then add 50μL 1% rabbit erythrocyte to each well and mixed gently. The plate is incubated for 3 hours at room temperature. The results are shown in Figure 1. It was obvious that the minimal effective concentration of GAL12 is 3.125 μg/mL.

Figure 2. The hemagglutination assay of GAL12 in V- bottom shaped 96-well microtiter plate.