Genetically encoded tags for direct synthesis of EM-visible gold nanoparticles in cells.
Nat Methods
; 17(9): 937-946, 2020 09.
Article
en En
| MEDLINE
| ID: mdl-32778831
ABSTRACT
Genetically encoded tags for single-molecule imaging in electron microscopy (EM) are long-awaited. Here, we report an approach for directly synthesizing EM-visible gold nanoparticles (AuNPs) on cysteine-rich tags for single-molecule visualization in cells. We first uncovered an auto-nucleation suppression mechanism that allows specific synthesis of AuNPs on isolated tags. Next, we exploited this mechanism to develop approaches for single-molecule detection of proteins in prokaryotic cells and achieved an unprecedented labeling efficiency. We then expanded it to more complicated eukaryotic cells and successfully detected the proteins targeted to various organelles, including the membranes of endoplasmic reticulum (ER) and nuclear envelope, ER lumen, nuclear pores, spindle pole bodies and mitochondrial matrices. We further implemented cysteine-rich tag-antibody fusion proteins as new immuno-EM probes. Thus, our approaches should allow biologists to address a wide range of biological questions at the single-molecule level in cellular ultrastructural contexts.
Texto completo:
1
Base de datos:
MEDLINE
Asunto principal:
Microscopía Electrónica
/
Nanopartículas del Metal
/
Oro
Límite:
Humans
Idioma:
En
Revista:
Nat Methods
Asunto de la revista:
TECNICAS E PROCEDIMENTOS DE LABORATORIO
Año:
2020
Tipo del documento:
Article
País de afiliación:
China