Dynamic Measurement of Endosome-Lysosome Fusion in Neurons Using High-Content Imaging.
Methods Mol Biol
; 2683: 201-212, 2023.
Article
em En
| MEDLINE
| ID: mdl-37300777
ABSTRACT
Endocytosis is a dynamic cellular process that actively transports particles into a cell. Late endosome fusion with the lysosome is a crucial step in the delivery of newly synthesized lysosomal proteins and endocytosed cargo for degradation. Disturbing this step in neurons is associated with neurological disorders. Thus, studying endosome-lysosome fusion in neurons will provide new insight into the mechanisms of these diseases and open new possibilities for therapeutic treatment. However, measuring endosome-lysosome fusion is challenging and time consuming, which limits the research in this area. Here we developed a high throughput method using pH-insensitive dye-conjugated dextrans and the Opera Phenix® High Content Screening System. By using this method, we successfully separated endosomes and lysosomes in neurons, and time-lapse images were collected to capture endosome-lysosome fusion events in hundreds of cells. Both assay set-up and analysis can be completed in an expeditious and efficient manner.
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Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Endossomos
/
Lisossomos
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article