Intelectin displays carbohydrate binding capacity and has been demonstrated to agglutinate bacteria, suggesting its role in innate immunity. It has also been linked to many pathogenic conditions in human. After reporting two amphioxus orthologs and the zebrafish intelectin 2 (zITLN2), here we cloned and characterized zebrafish intelectin 1 (zITLN1). Like zITLN2, zITLN1 also contains a conserved fibrinogen-related domain (FReD) and a unique intelectin domain (ITLN-D), expresses in all the tissues tested, with the highest level in intestine, and responds to bacterial challenge in acute phase. We also expressed zITLN1 in E. coli system, and purified recombinant zITLN1 could agglutinate both Gram-positive and Gram-negative bacteria in a calcium dependent manner. Its ability to agglutinate Gram-positive bacteria is stronger than that to Gram-negative bacteria whereas zITLN2 did not show such preference. This is probably due to the fact that recombinant zITLN1 could bind peptidoglycan (PGN) with a higher degree to lipopolysaccharide (LPS). Our results of zITLN1 provided new insight into the evolution and function of the intelectin family.