Fig. 3.

Preparation of an in situ TEM lamella across a large portion of the cell’s cytoplasm. (A) TEM micrographs stitched together to obtain an overview image of the lamella. On the left side the border region between ice and cell membrane can be recognized (black arrowhead). Within the lamella, the nuclear envelope (white arrowhead) can be clearly discerned, separating the nucleoplasm from the cytoplasm. The white arrow indicates the milling direction and highlights a prominent curtaining streak across the lamella. (B) Corresponding lamella imaged at lower magnification after recording of a tomographic tilt series. The dashed square regions denote the areas for tomographic exposure and focus, which are arranged along the tilt axis (for the corresponding tomographic reconstruction see Fig. 4). Some frost particles can be discerned across the lamella and on its sides (white asterisks in A and B). The framed region in (A) corresponds to the tomographic exposure region shown in (B). [Scale bars, (A) 1 μm, (B) 2 μm].