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Purpose: The aim of this retrospective study was to investigate the use of a radiopaque tissue fiducial marker (TFM) in the treatment of prostate cancer patients who undergo post-prostatectomy radiotherapy (PPRT). TFM safety, its role and benefit in quantifying the set-up uncertainties in patients undergoing PPRT image-guided radiotherapy were assessed. Purpose: The aim of this retrospective study was to investigate the use of a radiopaque tissue fiducial marker (TFM) in the treatment of prostate cancer patients who undergo post-prostatectomy radiotherapy (PPRT). TFM safety, its role and benefit in quantifying the set-up uncertainties in patients undergoing PPRT image-guided radiotherapy were assessed. Materials and Methods: A total of 45 consecutive PPRT patients underwent transperineal implantation of TFM at the level of vesicourethral anastomosis in the retrovesical tissue prior to intensity-modulated radiotherapy. Prostate bed motion was calculated by measuring the position of the TFM relative to the pelvic bony anatomy on daily cone-beam computed tomography. The stability and visibility of the TFM were assessed in the initial 10 patients. Results: No postoperative complications were recorded. A total of 3,500 images were analysed. The calculated prostate bed motion for bony landmark matching relative to TFM were 2.25 mm in the left-right, 5.89 mm in the superior-inferior, and 6.59 mm in the anterior-posterior directions. A significant 36% reduction in the mean volume of rectum receiving 70 Gy (rV70) was achieved for a uniform planning target volume (PTV) margin of 7 mm compared with the Australian and New Zealand Faculty of Radiation Oncology Genito-Urinary Group recommended PTV margin of 10 mm. Conclusion: The use of TFM was safe and can potentially eliminate set-up errors associated with bony landmark matching, thereby allowing for tighter PTV margins and a consequent favourable reduction in dose delivered to the bladder and rectum, with potential improvements in toxicities. [ABSTRACT FROM AUTHOR]