Purpose To investigate the effects of laser suture lysis (LSL) on filtration openings after trabeculectomy. Methods Prospective study analyzing the changes in the location and width of filtration openings, fluid cavity height, total bleb height, bleb wall thickness, and bleb wall intensity before and after LSL using threedimensional anterior segment optical coherence tomography (3D AS-OCT). Results Fourteen patients had clear scleral flap image analysis. As five patients underwent LSL twice and two patients underwent LSL thrice, 23 comparison studies were possible. After LSL the intraocular pressure (IOP) decreased (P = 0.0015) from 20.5 ± 5.3 to 14.9 ± 6.4 mm Hg, and the fluid cavity height increased significantly from 0.2 ± 0.2 mm to 0.3 ± 0.1 mm (P = 0.0094). Other bleb parameters were not significantly different when comparing before and after LSL. When the IOP reduction ratio was 425% following LSL, the width of the filtration openings on the LSL side, the total bleb height, and the fluid cavity height increased (P = 0.0273, 0.0342, and 0.0024, respectively). In multiple regression analysis the changes in fluid cavity height, the wall thickness, the wall intensity, and the width of the filtration opening were positively associated with the IOP reduction rate (P = 0.0428, 0.0226, 0.0420, and 0.0356, respectively). Conclusions 3D AS-OCT allowed a detailed examination of the internal morphology of filtration blebs and openings before and after LSL. The changes in the internal morphology were closely associated with the success of LSL to decrease IOP.