Real-time sentinel lymph node biopsy guidance using combined ultrasound, photoacoustic, fluorescence imaging: in vivo proof-of-principle and validation with nodal obstruction.

Precise sentinel lymph node (SLN) identification is crucial not only for accurate diagnosis of micro-metastases at an early stage of cancer progression but also for reducing the number of SLN biopsies (SLNB) to minimize their severe side effects. Furthermore, it is desirable that an SLNB guidance should be as safe as possible in routine clinical use. Although there are currently various SLNB guidance methods for pre-operative or intra-operative assessment, none are ideal. We propose a real-time SLNB guidance method using contrast-enhanced tri-modal images (i.e., ultrasound, photoacoustic, and fluorescence) acquired by a recently developed hand-held tri-modal probe. The major advantage of tri-modal imaging is demonstrated here through an in vivo study of the technically-difficult case of nodal obstruction that frequently leads to false-negative results in patients. The results in a tumor model in rabbits and normal controls showed that tri-modal imaging is capable of clearly identifying obstructed SLNs and of indicating their metastatic involvement. Based on these findings, we propose an SLNB protocol to help surgeons take full advantage of the complementary information obtained from tri-modal imaging, including for pre-operative localization, intra-operative biopsy guidance and post-operative analysis.