The aim of this study was to investigate the effects of tooth type (molar/premolar), tooth surface (buccal/lingual), smear layer or no smear layer, storage of specimens in deionised water, mode and speed of agitation, and rinsing method between cycles on mean step height loss and Knoop microhardness (KHN) change. Polished human enamel specimens embedded in acrylic resin were prepared from sound permanent molar and premolar tooth surfaces. A 0.3% (pH 3.2) solution of citric acid was used to erode the specimens, in a cycling procedure, consisting of 10 min immersion followed by rinsing in deionised water for 30 s with a spray bottle, for 5 cycles. The specimens were analysed with a non-contact white light profilometer and KHN. Molar teeth (148.99 ± 24.49 KHN) and buccal surfaces (155.62 ± 30.35 KHN) produced significantly less microhardness change compared to premolar (186.40 ± 20.74 KHN) and lingual surfaces (179.76 ± 23.21 KHN; p < 0001). The effect of storage and rinsing showed little difference in mean step height loss (<1 µm) and microhardness change. With no smear layer a significantly lower mean step height loss and microhardness change (p < 0.001) was observed. Agitation was performed with Orbital, Gyro and See-Saw rockers at 30, 40, 60, and 70 rpm. The mean step height loss was largest for See-Saw at 70 rpm (11.73 ± 0.91 μm) and lowest for Orbital at 30 rpm (2.76 ± 1.12 μm). A statistical difference was found between all types of agitation and speeds (p < 0.001). In conclusion, this study has shown that the variables investigated here have a significant impact on the measurable outcome, highlighting the importance of accurate and detailed method sections.

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