Type of presentation: Invited

LS-12-IN-6066 Correlative video-light-electron microscopy: development, impact and perspectives.

Rizzo R.1, Parashuraman R.1, Luini A.1
1Institute of Protein Biochemistry, National Research Council (CNR), Via Castellino 111, Naples, Italy
a.luini@ibp.cnr.it

GFP-based video microscopy provides profound insight into biological processes by generating information on the ‘history’, or dynamics, of the cellular structures involved in such processes in live cells. A crucial limitation of this approach, however, is that many important structures may not be resolved by light microscopy. Like more recent super-resolution techniques, Correlative video-Light-Electron Microscopy (CLEM) was developed to overcome this limitation. CLEM integrates GFP-based video and electron microscopy through a series of ancillary techniques such as proper fixation, hybrid labeling and retracing, and so provides the needed resolution. Another key characteristic of CLEM is that, by virtue of its electron microscopy component, it defines the context of the object of interest through the staining of bulk membranes and proteins and hence delineates all of the organelles and structure that surround or contact the structures of interest. This feature has no equivalent in light-based super-resolution techniques, which can only detect the fluorescent molecule used as marker. CLEM ‘multiplies’ the power of video microscopy, and is having an impact in several areas cell and developmental biology. Future developments of CLEM will, in our view, derive from combining again information deriving from GFP-based video microscopy (with its unique power to provide insight into the spatial-temporal organization of a process), with other types of information based on different imaging technology. In particular, correlating GFP-based video microscopy with the detection of large multi-molecular complexes, or with the localization of individual lipid species, will have significant impact in cell biology. Techniques that are suitable to achieve these aims are being developed. The potential, limitations and perspectives of correlative approaches aimed at integrating the unique insight generated by video microscopy with information from different forms of imaging will be discussed.