Abstract: This study aimed to investigate the characteristics of Toxoplasma gondii surface antigen 1 (SAG1) adhering to heparan sulfate on the host cell surface and the roles of SAG1 during T. gondii invasion in host cells. The recombinant plasmid pGEX-SAG1 was obtained by extracting the DNA of Toxoplasma gondii ME49 strain, amplifying the SAG1 gene fragment by PCR and cloning it into the prokaryotic expression vector pGEX-4T-1. The obtained recombinant plasmid pGEX-SAG1 was transformed into Escherichia coli BL21 CodonPlus-RIPL and induced by IPTG to express the recombinant protein GST-SAG1. The purified recombinant protein GST-SAG1 was used to analyze the heparin-binding characteristics, and the activity of SAG1 adherence to host cells was detected by IFA and Western blot. The heparin inhibition assays were used to further analyze the effects that the exogenous heparin blocked SAG1 adhering to the host cell surface in vitro and in vivo. The results showed that the recombinant expression vector pGEX-SAG1 was successfully constructed and then GST-SAG1 protein was expressed and purified. Heparin binding and competitive inhibition experiments showed that GST-SAG1 protein could bind to heparin, and the binding could be inhibited by heparin in a concentration-dependent manner. IFA and Western blot analysis indicated that SAG1 protein could adhere to the surface of Vero and HEK293A cells. The heparin inhibition assays showed the exogenous heparin could competitively inhibit SAG1 binding to heparan sulfate on the host cell surface to block T. gondii invasion. These results suggest T. gondii SAG1 involved in adhering to heparan sulfate on the host cell surface, contributing to invading host cells. This finding further reveals the interaction between SAG1, an important factor of T. gondii invasion, and heparan sulfate on the host cell surface during T. gondii invasion, it also lays the foundation for elucidating the molecular mechanism of heparin in the process of T. gondii invasion.