Introduction: Dentine hypersensitivity is a dental problem characterized by short, sharp pain araising from exposed dentine in response to stimuli, typically thermal, tactile, or chemical. One of the treatments for hypersensitive den-tine is by applying desensitizing agent such as hydroxyapatite which can occlude dentinal tubules. The fish bone is a natural material that can be used to synthesize hydroxyapatite. This study aimed to determine the effect of hydroxy-apatite from bone waste of Oreochromis niloticus, with a local name, tilapia, as a desensitizing agent on dentinal tubule occlusion. Methods: Hydroxyapatite powder was prepared using the precipitation method and sintered at 9000C for 5 hours. A 25% hydroxyapatite solution was obtained by mixing 25 grams of hydroxyapatite powder with 100 ml of distilled water. Dentine samples were prepared from twelve premolars cut longitudinally to separate the buccal and palatal sections, then the enamel layers on occlusal and buccal area were removed until the denti-no enamel junction. Dentine samples were etched with 6% citric acid to open dentine tubule. Occluded dentinal tubules were observed by measuring microstructure microscope for pretest and posttest after application of 25% hydroxyapatite solution (Group A) and distilled water (Group B). Data were analyzed statistically using paired t t-test and independent t-test. Results: The dentinal tubules occlusion in the pretest hydroxyapatite group was 41.17% and post test 95.94%, while in the pretest distilled water group was 35.90% and posttest 42.19%. The mean differences of dentinal tubule occlusion between pretest and posttest in groups A and B were significant, and also between the two groups. Conclusion: The present study showed that application of 25% hydroxyapatite solution from Oreochromis niloticus is effective to occlude dentinal tubules, so it could be considered for a dentine hypersensitivity treatment.