Desensitization of teeth after cavity preparation has been recommended in an attempt to avoid post-operative sensitivity. However, there is concern regarding application effect of desensitizing agent on shear bond strength of the adhesive system used. The purpose of our study was to compare the shear bond strength of adhesive system in two different dentin surface treatments, with and without desensitizing agent. Sixteen extracted human premolars were sectioned off at the coronal portion to expose the flat dentin surfaces. The surfaces were finished using 600 Grit Wet Silicon Carbide abrasive papers. The premolars were randomly assigned to two groups: control and treated with MS Coat desensitizing agent. The desensitizer was applied according to manufacturer’s instruction. Resin composite was bonded to each dentin surface using Prime & Bond ® adhesive system. The composite resin was debonded by shear stress. Mann-Whitney Test was used in statistical analysis. Our result showed that application of MS Coat desensitizing agent on dentin surface had significantly reduced the shear bond strength of the adhesive system used (z = - 0.14, p < 0.05). Thus, we conclude that shear bond strength of Prime & Bond ® NT (Dentsply, USA) adhesive system will be reduced if dentin surface is treated with MS Coat (Sun Medical, Japan) desensitizing agent.