An optimized protocol for immuno-electron microscopy of endogenous LC3.

ABSTRACT: MAP1LC3/LC3 (microtubule associated protein 1 light chain 3) is widely used as marker of autophagic compartments at different stages of maturation. Electron microscopy (EM) combined with immunolabeling is the only technique that can reveal the ultrastructural identity of LC3-labeled compartments. However, immuno-EM of endogenous LC3 proteins has proven difficult. Here, we test a panel of commercially available antibodies and apply different labeling conditions to present an optimized procedure for LC3 immuno-EM. Using ultrathin cryosections and protein A-colloidal gold or gold enhancement labeling, we localize endogenous LC3 in starved cells or tissues in the presence or absence of the proton pump inhibitor bafilomycin A₁. We localize LC3 to early and late stage autophagic compa

SUBMITTER: De Maziere A 

PROVIDER: S-EPMC9673964 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature