Carcinoembryonic antigen (CEA) which are characterized as members of the CD66 cluster of differentiation have highly related glycoproteins involved in cell adhesion. It also can be used as a tumor marker in clinical tests. CEA are glycosyl phosphatidyl inositol (GPI) cell-surface-anchored glycoproteins whose specialized sialofucosylated glycoforms serve as functional colon carcinoma L-selectin and E-selectin ligands. Besides, Galectin 4 (GAL4) has been identified as an interactor of CEA, thus a binding ELISA assay was conducted to detect the interaction of recombinant human CEA and recombinant human GLA4. Briefly, CEA were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100uL were then transferred to GLA4-coated microtiter wells and incubated for 2h at 37°C. Wells were washed with PBST and incubated for 1h with anti-CEA pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37°C. Finally, add 50µL stop solution to the wells and read at 450nm immediately. The binding activity of CEA and GLA4 was shown in Figure 1, and this effect was in a dose dependent manner.