This study evaluated the effect of repeated etching cycles on resin infiltrant penetration. Enamel samples measuring 4 × 4 × 3 mm 3 were obtained from the facial aspect of 50 extracted bovine teeth. Samples were immersed in a demineralization solution for 21 days to create artificial lesions and divided into five equal groups (n = 10). A 15% hydrochloric acid gel was administered to each group. The acid etching application time differed between groups: Group 1; 2 min, Group 2; 2 × 2 min, Group 3; 3 × 2 min, Group 4; 4 × 2 min, and Group 5; 5 × 2 min. Resin infiltration was visualized using a confocal laser scanning microscopy. The lesion, penetration and erosion depth (µm) were calculated, and data were statistically analyzed. The highest penetration depth (75.59 ± 9.42 µm) was seen in Group 5, followed by Groups 4, 3, 2 and 1. However, there were no statistically significant differences in the penetration depths between Groups 4 and 5 and between Groups 2, 3 and 4 (p > 0.05). In conclusion, a repeated etching cycle enhanced resin infiltrant penetration.