Three reasons explain why most of the critical events driving normal and pathological scenarios had been less investigated: they occur rarely in space and time, they are highly dynamic, they differ when studied in situ in an entire living organism. Metastasis is the primary cause for cancer-related mortality, but its mechanisms remain to be elucidated. Intravital imaging has opened the door to in vivo functional imaging in animal models of cancer, however it is limited in resolution. Ultrastructural analysis of tumor metastasis in vivo has so far been hindered by the limited field of view of the electron microscope, making it difficult to retrieve volumes of interest in complex tissues. We recently developed a multimodal correlative approach allowing us to rapidly and accurately combine functional in vivo imaging with high-resolution ultrastructural analysis of tumor cells in a relevant pathological context. The multimodal correlative approach that we propose here combines two-photon excitation microscopy (2PEM), microscopic X-ray computed tomography (microCT) and three-dimensional electron microscopy (3DEM). It enables a rapid and accurate correlation of functional imaging to high-resolution ultrastructural analysis of tumor cells in a relevant pathological context. As an example, we are now capable of providing high- and isotropic (8nm) resolution imaging of single metastasizing tumor cells previously imaged in the process of extravasation in the living mouse brain. This reliable and versatile workflow offers access to ultrastructural details of metastatic cells with an unprecedented throughput opening to crucial and unparalleled insights into the mechanisms of tumor invasion, extravasation and metastasis in vivo.

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EMC Abstracts - https://emc-proceedings.com/abstract/tracking-tumor-metastasis-in-vivo-at-high-resolution/