Background : This study was designed to explore the feasibility of replacing the conventional peri-/intratumoural ultrasound (US)-guided technetium-99m albumin nanocolloid (99mTc-nanocolloid) administration by an injection of the same tracer guided by a freehand single-photon emission computed tomography (SPECT) device in patients with non-palpable breast cancer with an iodine-125 (125I) seed as tumour marker, who are scheduled for a sentinel lymph node biopsy (SLNB). This approach aimed to decrease the workload of the radiology department, avoiding a second US-guided procedure. ; Methods : In ten patients, the implanted 125I seed was primarily localised using freehand SPECT and subsequently verified by conventional US in order to inject the 99mTc-nanocolloid. The following 34 patients were injected using only freehand SPECT localisation. In these patients, additional SPECT/CT was acquired to measure the distance between the 99mTc-nanocolloid injection depot and the 125I seed. In retrospect, a group of 21 patients with US-guided 99mTc-nanocolloid administrations was included as a control group. ; Results : The depth difference measured by US and freehand SPECT in ten patients was 1.6 ± 1.6 mm. In the following 36 125I seeds (34 patients), the average difference between the 125I seed and the centre of the 99mTc-nanocolloid injection depot was 10.9 ± 6.8 mm. In the retrospective study, the average distance between the 125I seed and the centre of the 99mTc-nanocolloid injection depot as measured in SPECT/CT was 9.7 ± 6.5 mm and was not significantly different compared to the freehand SPECT-guided group (two-sample Student's t test, p = 0.52). ; Conclusion : We conclude that using freehand SPECT for 99mTc-nanocolloid administration in patients with non-palpable breast cancer with previously implanted 125I seed is feasible. This technique may improve daily clinical logistics, reducing the workload of the radiology department.