To evaluate (1) the effect of a desensitizing paste containing 8% arginine and calcium carbonate on the microtensile bond strength between dentin and etch-and-rinse adhesive systems; and (2) to examine the dentin tubules occlusion quantitatively. 48 freshly extracted intact human mandibular third molars were divided randomly into three groups. The mid-coronal dentin of each tooth was exposed and treated. Group A: no treatment; Group B: specimens were polished with a desensitizing paste containing 8% arginine and calcium carbonate using a rotary cup operating at a low speed for 3 seconds, followed by an additional duration of 3 seconds (total operation time of 6 seconds), according to the manufacturer's instructions; Group C: specimens were handled in the same way with the exception of an increased operation time of 9 seconds, twice (total operation time of 18 seconds). Each group was randomly divided into two subgroups in order to evaluate the effectiveness of two different adhesive agents. A two-step etch-and-rinse adhesive agent (Adper SingleBond 2) and a three-step etch-and-rinse adhesive agent (Adper ScotchBond Multi-purpose) were applied to dentin surfaces. Then, microtensile bond strengths of the six subgroups were tested. Dentin surfaces were analyzed using field-emission scanning electron microscopy (FESEM) and laser scanning confocal microscopy (LSCM). There was no significant difference in microtensile bond strength between the control group and the experimental groups treated with the 8% arginine and calcium carbonate desensitizing paste during the application of etch-and-rinse adhesives. Both FESEM and LSCM showed that the desensitizing paste occluded dentin tubules effectively.