Surface Guided Radiotherapy (SGRT) is a relatively new technique for positioning patients and for monitoring patient movement during treatment. SGRT is completely non-invasive since it uses visible light for determining the position of the patient surface. A reduction in daily imaging for patient setup is possible if the accuracy of SGRT is comparable to imaging. It allows for monitoring of intrafraction motion and the radiation beam can be held beyond a certain threshold resulting in a more accurate irradiation. The purpose of this study was to investigate setup uncertainty and the intrafraction motion in non-gated whole breast cancer radiotherapy treatment using an integrated implementation of AlignRT (OSMS) system as SGRT. In initial setup, SGRT was compared to three-point setup using tattoos on the patient and orthogonal kV imaging. For the investigation of intrafraction motion, OSMS monitored the patient with six degrees of freedom during treatment. Using three-point setup resulted in a setup root-mean-square error from the isocenter of 5.4 mm. This was improved to 4.2 mm using OSMS. For the translational directions, OSMS showed improvements in the lateral direction (P = 0.0009, Wilcoxon rank-sum), but for the longitudinal direction and rotation it was not possible to show improvements (P = 0.96 and P = 0.46, respectively). The vertical direction proved more accurate for three-point setup than OSMS (P = 0.000004). Intrafraction motion was very limited with a translational median of 1.1 mm from the isocenter. While OSMS showed marked improvements over laser and tattoo setup, the system did not prove accurate enough to replace the daily orthogonal kV images aligned to bony anatomy.