This study evaluated the long-term microleakage of access cavities conditioned with phosphoric acid and deproteinizing agents for root-canal-treated teeth using fluid filtration and microscopical analysis. Occlusal surfaces of extracted human mandibular premolars (N=90) were removed leaving a 4mm coronal length from the cemento-enamel junction. After root canal treatment, the specimens were randomly divided into four experimental groups (n=21) and the remaining teeth were used for positive control group (n=6): SB: 35% H3PO4+Adper Single Bond 2; SSB: 35% H3PO4+5.25% NaOCl+10% Sodium ascorbate (C6H7NaO6) + Adper Single Bond 2; XP: 35% H3PO4+XP Bond; SXP: 35% H(3)PO(4+)5.25% NaOCl+10% Sodium ascorbate+XP Bond. All cavities were restored with a resin composite (Filtek Z250). After removing the root filling from the apical side, teeth were subjected to fluid filtration test for 1week, 6 and 12months followed by x2500 thermocycling after 1st week and 6th months each. Data were analyzed using one-way ANOVA and Dunnet T3 tests (alpha=0.05). SEM analyses were carried out after each microleakage evaluation in two random teeth from all groups. Microleakage values in groups SB and XP presented no significant difference in any of the evaluated period (p>0.05). Microleakage results of SXP (0.01665) group showed significant difference compared to XP (0.03377) and SB (0.03049) groups after 12months. SSB group (0.00901) showed significantly less microleakage among all other groups (0.01665-0.03377) (p<0.05). Prior to endodontic treatment, in access cavities, acid etching with 35% H3PO4 followed by the application of NaOCl and sodium ascorbate completely destroyed the collagen layer, reducing the microleakage and resin-dentin interface degradation up to 12months.