Objectives This laboratory study aimed to: a) compare the fluoride dose-response of different caries lesions created in human and bovine enamel (HE/BE) under pH-cycling conditions, and b) investigate the suitability of Knoop and Vickers surface microhardness (K-SMH/V-SMH) in comparison to transverse microradiography (TMR) to investigate lesion de-and remineralization.Materials and Methods Caries lesions were formed using three different protocols (Carbopol, hydroxyethylcellulose-HEC, methylcellulose-MeC) and assigned to 24 groups using V-SMH, based on a 2 (enamel types)×3 (lesion types)×4 (fluoride concentrations used during pH-cycling-simulating 0/250/1100/2800 ppm F as sodium fluoride dentifrices) factorial design. Changes in mineral content and structural integrity of lesions were determined before and after pH-cycling. Data were analyzed using three-way ANOVA.Results BE was more prone to demineralization than HE. Both enamel types showed similar responses to fluoride with BE showing more remineralization (as change in integrated mineral loss and lesion depth reduction), although differences between tissues were already present at lesion baseline. Carbopol and MeC lesions responded well to fluoride, whereas HEC lesions were almost inert. K-and V-SMH correlated well with each other and with the integrated mineral loss data, although better correlations were found for HE than for BE, and for MeC than for Carbopol lesions. Hardness data for HEC lesions correlated only with surface zone mineral density data.Conclusion BE is a suitable surrogate for HE under pH-cycling conditions. Clinical Relevance The in vitro modelling of dental caries is complex and requires knowledge of lesion behavior, analytical techniques and employed hard tissues.