Send to

Choose Destination
PLoS One. 2014 Dec 5;9(12):e114448. doi: 10.1371/journal.pone.0114448. eCollection 2014.

Correlating intravital multi-photon microscopy to 3D electron microscopy of invading tumor cells using anatomical reference points.

Author information

European Molecular Biology Laboratory Heidelberg, Heidelberg, 69117, Germany.
Inserm U1109, MN3T, Strasbourg, F-67200, France; Université de Strasbourg, Strasbourg, F-67000, France; LabEx Medalis, Université de Strasbourg, Strasbourg, F-67000, France; Fédération de Médecine Translationnelle de Strasbourg (FMTS), Strasbourg, F-67000, France.
Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America.


Correlative microscopy combines the advantages of both light and electron microscopy to enable imaging of rare and transient events at high resolution. Performing correlative microscopy in complex and bulky samples such as an entire living organism is a time-consuming and error-prone task. Here, we investigate correlative methods that rely on the use of artificial and endogenous structural features of the sample as reference points for correlating intravital fluorescence microscopy and electron microscopy. To investigate tumor cell behavior in vivo with ultrastructural accuracy, a reliable approach is needed to retrieve single tumor cells imaged deep within the tissue. For this purpose, fluorescently labeled tumor cells were subcutaneously injected into a mouse ear and imaged using two-photon-excitation microscopy. Using near-infrared branding, the position of the imaged area within the sample was labeled at the skin level, allowing for its precise recollection. Following sample preparation for electron microscopy, concerted usage of the artificial branding and anatomical landmarks enables targeting and approaching the cells of interest while serial sectioning through the specimen. We describe here three procedures showing how three-dimensional (3D) mapping of structural features in the tissue can be exploited to accurately correlate between the two imaging modalities, without having to rely on the use of artificially introduced markers of the region of interest. The methods employed here facilitate the link between intravital and nanoscale imaging of invasive tumor cells, enabling correlating function to structure in the study of tumor invasion and metastasis.

[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Public Library of Science Icon for PubMed Central
Loading ...
Support Center