Objectives The aim of the present study involved comparing the tubular occluding properties of three varnishes, two Clinpro™ varnishes and a Colgate® Duraphat® Varnish. Method Nine caries-free premolar dentine discs were prepared and treated with the three varnishes and subsequently observed under SEM at 0o and 90o angle to assess their ability to cover the dentine surface. The tubule occluding properties were measured using a modified Pashley cell hydraulic conductance model. A further 9 caries-free extracted molars were sectioned into 500μ thick dentine discs. The fluid flow rate was assessed after a) immersing the discs in 6% citric acid for 2 minutes, b) treating the dentine disc with the experimental varnishes and c) subjecting the treated discs to an acid challenge (6% citric acid for 2 minutes). Results SEM investigation showed uniform occlusion of the dentinal tubules, with varying depths of penetration. Hydraulic conductance tests showed no statistically significant differences in the fluid flow rate (expressed as percentages) when all the three varnishes were compared at different stages of treatment (p = 0.33). However, after subjecting the discs to an acid challenge, there was a statistically significant increase in the fluid flow rate with the Colgate® Duraphat® Varnish treated discs, whereas the ClinproTM White Varnish and Clinpro™ XT Varnish discs showed no statistically significant differences (p = 0.99 and p= 0.83 respectively). Conclusions All the tested varnishes (Colgate® Duraphat® Varnish, ClinproTM White Varnish and ClinproTM XT varnish) were effective in blocking the dentinal tubules as demonstrated in this in vitro study. However, the tricalcium phosphate varnishes (ClinproTM) were more resistant to an in vitro acid challenge compared to the sodium fluoride varnish (Colgate® Duraphat®)