Monitoring the Localization of MAP1LC3B by Indirect Immunofluorescence

Abstract

The autophagy protein MAP1LC3B (microtubule-associated proteins 1A/1B light chain 3B, hereafter referred to as LC3B), which is one of several mammalian homologs of yeast Atg8, is one of the most popular markers for autophagosome formation because its distribution changes from cytosolic/diffuse to punctate upon the induction of autophagy. In many settings, plasmids encoding fluorescently tagged LC3B are introduced into cells, and the subsequent autophagy response is monitored. However, for a variety of reasons, it would be desirable also to have a protocol to monitor the localization of endogenous LC3B under various conditions. This protocol provides such a methodology for the staining of endogenous LC3B by indirect immunofluorescence, such that autophagy responses can be monitored in mammalian cells.