Background and purpose: To determine the uncertainties in planned dose associated with catheter and organ movement during 48 hours of stepping source prostate brachytherapy. Material and methods: Pulsed-dose rate (PDR) prostate brachytherapy as a boost is given in 24 pulses every 2 hours, making the total treatment last 48 hours. The entire treatment is based on one plan, created on the planning CT (CT1). Two follow-up CTs (CT2 and CT3) were acquired; halfway through the treatment and at the end of treatment. On these repeat scans the catheters were reconstructed and PTV and OARs were delineated. The original treatment plan was calculated on the repeat CTs. Target coverage V 100%, D90, dose to 2 cm3 (D2cm3) of the rectum and bladder and dose to 0.1 cm3 of the urethra were recorded from the recalculated DVHs. Results: On the two repeat CTs the V 100% decreased -1.5% and -2.3% as compared to the planning CT. For the rectum D2cm3, the average increase was 14.8% (CT1-CT2) and 17.3% (CT1-CT3). Increase in bladder D2cm3 was on average 23.1% (CT1-CT2) and 24.8% (CT1-CT3). For the urethra D0.1cm3 an average decrease of -2% (CT1-CT2) and -3.2% (CT2-CT3) was observed. Conclusions: Changes in target coverage during treatment were small and considered clinically irrelevant. However, an overall increase in dose to the OARs was found as compared to the planned dose, which should be taken into account during treatment planning. © 2012 Elsevier Ireland Ltd. All rights reserved.