3D imaging of mammalian cells with ion-abrasion scanning electron microscopy.
Understanding the hierarchical organization of molecules and organelles within the interior of large eukaryotic cells is a challenge of fundamental interest in cell biology. We are using ion-abrasion scanning electron microscopy (IA-SEM) to visualize this hierarchical organization in an approach that combines focused ion-beam milling with scanning electron microscopy. Here, we extend our previous studies on imaging yeast cells to image subcellular architecture in human melanoma cells and melanocytes at resolutions as high as approximately 6 and approximately 20 nm in the directions parallel and perpendicular, respectively, to the direction of ion-beam milling. The 3D images demonstrate the striking spatial relationships between specific organelles such as mitochondria and membranes of the endoplasmic reticulum, and the distribution of unique cellular components such as melanosomes. We also show that 10nm-sized gold particles and quantum dot particles with 7 nm-sized cores can be detected in single cross-sectional images. IA-SEM is thus a useful tool for imaging large mammalian cells in their entirety at resolutions in the nanometer range.
Heymann
Jurgen A W
JA
Laboratory of Cell Biology, Center for Cancer Research National Cancer Institute, NIH, Building 50, Room 4306, Bethesda, MD 20892, USA.
Shi
Dan
D
Kim
Sang
S
Bliss
Donald
D
Milne
Jacqueline L S
JL
Subramaniam
Sriram
S
D045744:Cell Line, Tumor; D003584:Cytological Techniques / Q000379:methods; D005057:Eukaryotic Cells / Q000648:ultrastructure*; D006046:Gold; D006801:Humans; D021621:Imaging, Three-Dimensional / Q000379:methods*; D007425:Intracellular Membranes / Q000648:ultrastructure; D008544:Melanocytes / Q000648:ultrastructure; D008545:Melanoma / Q000648:ultrastructure; D020460:Melanosomes / Q000648:ultrastructure; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D046529:Microscopy, Electron, Transmission; D008928:Mitochondria / Q000648:ultrastructure; D015388:Organelles / Q000648:ultrastructure; D045663:Quantum Dots; D013194:Staining and Labeling
D006046:Gold