Three-dimensional (3D) printing technology is highly regarded in the field of dentistry. Three-dimensional printed resin restorations must undergo a washing process to remove residual resin on the surface after they have been manufactured. However, the effect of the use of different washing solutions and washing times on the biocompatibility of the resulting resin restorations is unclear. Therefore, we prepared 3D-printed denture teeth and crown and bridge resin, and then washed them with two washing solutions (isopropyl alcohol and tripropylene glycol monomethyl ether) using different time points (3, 5, 10, 15, 30, 60, and 90 min). After this, the cell viability, cytotoxicity, and status of human gingival fibroblasts were evaluated using confocal laser scanning. We also analyzed the flexural strength, flexural modulus, and surface SEM imaging. Increasing the washing time increased the cell viability and decreased the cytotoxicity ( < 0.001). Confocal laser scanning showed distinct differences in the morphology and number of fibroblasts. Increasing the washing time did not significantly affect the flexural strength and surface, but the flexural modulus of the 90 min washing group was 1.01 ± 0.21 GPa (mean ± standard deviation), which was lower than that of all the other groups and decreased as the washing time increased. This study confirmed that the washing time affected the biocompatibility and mechanical properties of 3D printed dental resins.