Objective: In an aim to investigate aspects of an Artificial Mouth System (AMS) to model the caries process in bovine enamel biofilm induced white spot enamel lesions (WSEL) were analyzed. Methods: Square shaped and polished bovine enamel slabs were covered with modeling wax and 2.5 mm diameter window was prepared to expose enamel surface at the center of each slab. Artificial biofilms were grown inside an AMS on the slabs using freshly cultured Streptococcus mutans (MT8148) suspended in phosphate buffered saline (PBS), heart infusion (HI) with 1% sucrose and PBS for 20, 30 and 40 hrs at 37°C. The severity of demineralization was quantified by a Quantitative Light-induced Fluorescence (QLF, Inspector, Netherlands), nano-mechanical property changes were studied by a Nanoindenter (ENT-1100, Elionix, Japan), fine-structural changes were observed by SEM after argon ion-etching. All experiments were repeated three times (n=5). Results: A circular WSEL could clearly be detected at the center of each slab by naked eyes, which was prominent after dehydration by mild air drying. QLF data showed that severity of demineralization occurred with the increase of time interval, after 40 hours both DF (23±2.9) and DQ (148.75±26.78) were significantly more compared to DF (10.14±2.3) and DQ (47.63±30.67) of 20 hours. Hardness measured by nanoindenter at the center of all lesions was significantly reduced from unaffected enamel, 40-hr sample showed about 95% reductions. Perpendicularly trimmed surface topography after argon ion-etching by SEM clearly visualized depth of demineralized WSEL, widened inter-enamel prism channels, dislodgement of prism crystals and perforated prism structure at outer most layers. Conclusion: An advanced in vitro model of biolfilm induced WSEL was developed that showed promises to be useful in studying enamel demineralization and remineralization employing QLF, nano-indenter and SEM. This study was supported by COE program for FRMDRTB at TMDU and Japan Monbukagaku Research Grant (16390544).