An antibody for analysis of autophagy induction.
Nat Methods
; 17(2): 232-239, 2020 02.
Article
en En
| MEDLINE
| ID: mdl-31768061
Autophagy is a degradative program that maintains cellular homeostasis. Autophagy defects have been described in numerous diseases. However, analysis of autophagy rates can be challenging, particularly in rare cell populations or in vivo, due to limitations in currently available tools for measuring autophagy induction. Here, we describe a method to monitor autophagy by measuring phosphorylation of the protein ATG16L1. We developed and characterized a monoclonal antibody that can detect phospho-ATG16L1 endogenously in mammalian cells. Importantly, phospho-ATG16L1 is only present on newly forming autophagosomes. Therefore, its levels are not affected by prolonged stress or late-stage autophagy blocks, which can confound autophagy analysis. Moreover, we show that ATG16L1 phosphorylation is a conserved signaling pathway activated by numerous autophagy-inducing stressors. The described antibody is suitable for western blot, immunofluorescence and immunohistochemistry, and measured phospho-ATG16L1 levels directly correspond to autophagy rates. Taken together, this phospho-antibody represents an exciting tool to study autophagy induction.
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Autofagia
/
Anticuerpos
Límite:
Animals
/
Humans
Idioma:
En
Revista:
Nat Methods
Asunto de la revista:
TECNICAS E PROCEDIMENTOS DE LABORATORIO
Año:
2020
Tipo del documento:
Article
País de afiliación:
Canadá