Figure 4.

Correlative light-electron microscopy of EDS. Transfected A375MM melanoma cells were plated on CELLocate coverslips coated with cross-linked fluorophore-conjugated matrix for 15 h, fixed, analyzed at the confocal microscope, and then processed for immunoelectron microscopy. The shown electron microscopy fields are boxed in yellow in the corresponding confocal images, arrows indicate invadopodial protrusions. The boxed area of a wtDyn2aa-GFP–transfected cell (A) was serially sectioned and observed at the EM. The movie serial.mov shows the complete serial section series. Three different serial sections (bottom, middle, and top) (B) show Dyn2-GFP labeling as identified with anti-GFP antibodies. The movie stack.mov shows the complete Z-stack rendering of the confocal image series, whereas C and D show two different confocal sections of a same cell (substrate level and middle section, respectively); fragments of partially degraded gelatin are clearly visible both in the confocal (D) and electron microscopy images (E, arrowheads). The boxed area of Dyn2^K44A-GFP–transfected cells (F) was observed at the electron microscope; arrows point to structures vaguely resembling EDS but devoid of invadopodia and gelatin fragments. H shows an electron micrograph of a Dyn2^ΔPRD-GFP–transfected cell. The drawing depicts a schematic reconstruction of EDS in relationship to the nucleus (N) and the Golgi apparatus (GA), ECM is indicated in red. Bar, 10 μm in confocal images, and 1 μm in EM images.