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Barsha Shrestha,1 Sheetal Maria Rajan,1 Sultan Aati,1,2 Emielda Yusiharni,3 Omar Kujan,1 Martin Saunders,4 Amr Fawzy1 1UWA Dental School, The University of Western Australia, Perth, WA, Australia; 2Dental Health Department, College of Applied Medical Sciences, King Saud University, Riyadh, Saudi Arabia; 3UWA XRD Facility, Material & Environmental Analysis Platform, The University of Western Australia, Perth, WA, Australia; 4Centre for Microscopy, Characterisation and Analysis, The University of Western Australia, Perth, WA, AustraliaCorrespondence: Amr Fawzy, UWA Dental School, The University of Western Australia, 17 Monash Avenue, Nedlands, WA, 6009, Australia, Tel +61894572434, Email amr.fawzy@uwa.edu.auBackground: Remineralization of dental enamel is an important intervention strategy for the treatment of demineralized lesions. Existing approaches have limitations such as failure to adequately reproduce both the ideal structural and mechanical properties of the native tooth. The ability of ultrasound to control and accelerate the crystallization processes has been widely reported. Therefore, a new approach was explored for in-vitro enamel remineralization involving the synergistic effect of high-intensity focused ultrasound (HIFU) coupled with calcium phosphate ion clusters (CPICs).Methods: The demineralized enamel was treated with CPICs, with or without subsequent HIFU exposure for different periods (2.5, 5, and 10 min). The specimens were characterized by scanning electron microscopy (SEM), atomic force microscopy (AFM), and Raman spectroscopy. The surface hardness and crystallographic properties of the treated specimens were evaluated using Vickers microhardness testing and X-ray diffraction (XRD), respectively.Results: SEM revealed distinct, organized, and well-defined prismatic structures, showing clear evidence of remineralization in the combined CPIC/HIFU treatment groups. AFM further revealed a decrease in the surface roughness values with increasing HIFU exposure time up to 5 min, reflecting the obliteration of interprismatic spaces created during demineralization. The characteristic Raman band at 960 cm− 1 associated with the inorganic phase of enamel dominated well in the HIFU-treated specimens. Importantly, microhardness testing further demonstrated that new mineral growth also recovered the mechanical properties of the enamel in the HIFU-exposed groups. Critical to our aspirations for developing this into a clinical process, these results were achieved in only 5 min.Conclusion: HIFU exposure can synergise and significantly accelerate in-vitro enamel remineralization process via calcium phosphate ion clusters. Therefore, this synergistic approach has the potential for use in future clinical interventions. Keywords: enamel remineralization, high intensity focused ultrasound, calcium phosphate ion cluster