We aimed to analyze the effects of printing parameters on characterization of three-dimensional (3D) printed bolus used in external beam radiotherapy. Two sets of measurements were performed to investigate the dosimetric and physical characterization of 3D-printed bolus at different printing parameters. In the first step, boluses were produced at different infill-percentages, infill-patterns and printing directions. Two-dimensional (2D) dose measurements were performed in Elekta Versa HD linear accelerator using 6 MV photon energy. Measured 2D dose maps for both printed and reference bolus materials were compared using the 2D gamma analysis method. Additionally, patient-specific bolus was produced with defined optimum printing parameters for anthropomorphic head and neck phantom. Then, point dose measurements were performed to evaluate the feasibility of printed bolus in clinical use. In the second step, physical measurements were carried out to evaluate the printing accuracy, the mean hounsfield unit (HU) value and the weight of 3D-printed boluses. According to our measurement, infill-percentage, infill-pattern and printing direction significantly changed the dosimetric and physical properties of the 3D-printed bolus independently. Maximum gamma passing rate at 1.5 and 5 cm depths were found as 93.8% and 98.8%, respectively, for 60% infill-percentage, sunglass fill infill-pattern and horizontal printing direction. The printing accuracy of the products was within 0.4 mm. Dosimetric and physical properties of the printed bolus material changed significantly with the selected printing parameters. Therefore, it is important to note that each combination of these printing parameters that will be used in the production of patient-specific bolus should be investigated separately. ? 2020 American Association of Medical Dosimetrists. Published by Elsevier Inc. All rights reserved.