High-quality ultrastructural preservation using cryofixation for 3D electron microscopy of genetically labeled tissues.
Elife
; 72018 05 11.
Article
en En
| MEDLINE
| ID: mdl-29749931
ABSTRACT
Electron microscopy (EM) offers unparalleled power to study cell substructures at the nanoscale. Cryofixation by high-pressure freezing offers optimal morphological preservation, as it captures cellular structures instantaneously in their near-native state. However, the applicability of cryofixation is limited by its incompatibility with diaminobenzidine labeling using genetic EM tags and the high-contrast en bloc staining required for serial block-face scanning electron microscopy (SBEM). In addition, it is challenging to perform correlated light and electron microscopy (CLEM) with cryofixed samples. Consequently, these powerful methods cannot be applied to address questions requiring optimal morphological preservation. Here, we developed an approach that overcomes these limitations; it enables genetically labeled, cryofixed samples to be characterized with SBEM and 3D CLEM. Our approach is broadly applicable, as demonstrated in cultured cells, Drosophila olfactory organ and mouse brain. This optimization exploits the potential of cryofixation, allowing for quality ultrastructural preservation for diverse EM applications.
Palabras clave
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Microscopía Electrónica de Rastreo
/
Criopreservación
Límite:
Animals
Idioma:
En
Revista:
Elife
Año:
2018
Tipo del documento:
Article
País de afiliación:
Estados Unidos