The present study aimed to assess the sealing ability of various repair materials in furcation perforations using dye penetration. A total of 80 human first molars from mandible with unbroken enamel surfaces, which were extracted completely for periodontal reasons, were chosen. All extracted teeth were kept in a sterile solution up until further use. Endo-access bur was used to prepare endodontic access cavity, and a high-speed long shank round bur was used to create a planned perforation on the floor of pulp. All the specimens were randomly grouped (20 teeth in each group): Group I: control; group II: Pro-Root MTA; group III: resin-modified glass-ionomer cement (RM GIC); and group IV: Biodentin. After placement of repair materials in perforated furcation, the chamber of each tooth was accessed with 2% methylene blue dye. Later, a diamond disk was used to split the teeth buccolingually. A stereomicroscope (10×) was used to visualize the highest dye penetration. The least penetration was seen in teeth repaired by Biodentin group (0.88 ± 0.02), next by Pro-Root MTA group (1.24 ± 0.13) and resin-modified glass ionomer cement group (3.62 ± 0.46). A statistically significant difference was seen among different perforation repair materials by analysis of variance. A statistically significant difference (p value < 0.05) was found between each group, excluding group II v/s group IV. This study concluded that reduced dye penetration in perforation repair along the furcations and improved sealing ability are shown by biodentine group in comparison to Pro-Root MTA and resin-modified glass-ionomer cement group. Root perforations along the furcation that develop due to the endodontic procedures have a remarkable damaging effect on prognosis and frequently result in loss of secondary periodontal attachment, thus resulting in early loss of the involved tooth. Therefore, the selection of biocompatible repair material aids to enhance the treatment prognosis.