Visualizing HIV-1 Assembly at the T-Cell Plasma Membrane Using Single-Molecule Localization Microscopy.
Methods Mol Biol
; 2807: 61-76, 2024.
Article
em En
| MEDLINE
| ID: mdl-38743221
ABSTRACT
The 20-year revolution in optical fluorescence microscopy, supported by the optimization of both spatial resolution and timely acquisition, allows the visualization of nanoscaled objects in cell biology. Currently, the use of a recent generation of super-resolution fluorescence microscope coupled with improved fluorescent probes gives the possibility to study the replicative cycle of viruses in living cells, at the single-virus particle or protein level. Here, we highlight the protocol for visualizing HIV-1 Gag assembly at the host T-cell plasma membrane using super-resolution light microscopy. Total internal reflection fluorescence microscopy (TIRF-M) coupled with single-molecule localization microscopy (SMLM) enables the detection and characterization of the assembly of viral proteins at the plasma membrane of infected host cells at the single protein level. Here, we describe the TIRF equipment, the T-cell culture for HIV-1, the sample preparation for single-molecule localization microscopies such as PALM and STORM, acquisition protocols, and Gag assembling cluster analysis.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Linfócitos T
/
Membrana Celular
/
HIV-1
/
Montagem de Vírus
/
Produtos do Gene gag do Vírus da Imunodeficiência Humana
/
Imagem Individual de Molécula
/
Microscopia de Fluorescência
Limite:
Humans
Idioma:
En
Ano de publicação:
2024
Tipo de documento:
Article