Background: Intraoperative lymph node mapping (LNM) is highly significant for many surgeries in patients with cancer. Many types of tracers are currently used, but the ideal method has not yet been identified. We aimed to identify a stable lymphatic drainage pathway in an animal model and compared the effects of quantum dots (QD), a new fluorescent tracer, with those of methylene blue in intraoperative LNM. Materials and methods: Indian ink (0.2 mL) was subcutaneously injected into the plantar metatarsal regions of six Sprague-Dawley rats. After 2 wk of incubation and subsequent dissection, the potentially stained LNs were examined pathologically to identify the lymphatic drainage pathway. After applying anesthesia, 0.1 mL methylene blue (2%) and QD (1 mg/mL) were injected into the plantar metatarsal regions of six rats for intraoperative LNM. The QD group was observed with a near-infrared imaging system, and the methylene blue group was directly observed. Drainages were recorded at 5, 10, 30, 60, and 120 min and at 1 d. Results: Two three-level drainage pathways, that is, a peripheral drainage (popliteal LNs, inguinal LNs, and axillary LNs) and a central drainage (popliteal lymph node [LN], iliac LN, and renal LN) pathways were identified. Both methylene blue and QD stained the sentinel lymph node (SLNs) quickly, but methylene blue was difficult to identify in the deep tissues and the LNs beyond the SLN. Furthermore, the blue-stained LNs remain dyed for only 2 h. In contrast, the QDs exhibited high target-to-background ratios in both the SLNs and the following LNs. Additionally, the fluorescence lasted from 5 min-1 d after injection. Conclusions: An ideal lymphatic drainage model was found. QDs are excellent tracers for intraoperative LNM compared with methylene blue. Near infrared fluorescent imaging is a promising LNM method for clinical practice. (C) 2014 Elsevier Inc. All rights reserved.