This study evaluated the biomineralization and dentinal tubules occlusion abilities of the carboxyl-terminated polyamidoamine dendrimer (PAMAM-COOH) on human demineralized dentin in vivo at different time points. Demineralization dentin model with and without treated with PAMAM-COOH were sutured to the interior side of the rat's cheeks, that was incubated in the rats' saliva for 2, 4 and 6 weeks respectively. Finally, the newly formed precipitates were characterized by SEM, EDS, XRD and microhardness test. The hydroxyapatite (HA) on the dentin treated with PAMAM-COOH were formed gradually with the time going by, and the regenerated HA has a similar crystal structure with natural dentin, whereas the crystallites did not exist on the control group. The microhardness of PAMAM-COOH-applied specimens had a significantly higher than those without application. These results suggest that the PAMAM-COOH promoted the biomineralization of demineralized dentin and displayed favourable effects on blocking the open dentinal tubules.