Introduction: First line therapy of dentin hypersensitivity typically involves the use of desensitizing toothpastes. Two such agents in the market are 5% Calcium Sodium Phosphosilicate (CSPS) and 8% Arginine Calcium Carbonate (ACC), which have unique mechanisms of blocking the dentinal tubules in a natural bio-mimetic way. So the purpose of this in vitro study was to evaluate and compare the efficacy of dentinal tubule occlusion using Scanning Electron Microscope (SEM) and dentinal permeability by using Stereomicroscope (SM) after the application of these agents. Methods: Total 60 samples were included in this study. 30 samples were used for SEM analysis and 30 others for SM analysis. Each of these 30 samples were again divided into three groups as: distilled water-Group-I, 5% CSPS-Group-II and 8% ACC-Group-III. The dentine surface morphology was evaluated using SEM after brushing and after acid treatment. Dentine permeability analysis using Evans blue dye was evaluated by using SM. Statistical analysis was done using one-way ANOVA and Chi-square test. Tukey post hoc test was used for multiple comparisons. Results: CSPS and ACC test groups showed statistically significant difference in dentinal tubule occlusion and in permeability reduction than the respective DW control (p<0.001). Comparing the mean permeability differences between different groups, after the treatment, CSPS group showed a higher permeability reduction and number of occluded tubules. Conclusion: 5% CSPS shows promise as a desensitizing agent, which not only occludes the dentinal tubules, but also reduces the tubule permeability and shows very good resistance to acid treatment than 8% ACC.