Methods to Detect Loss of Lysosomal Membrane Integrity

Abstract

Loss of lysosomal membrane integrity, often referred to as lysosomal membrane permeabilization (LMP), occurs in many instances of cell death either as an initiating or as an amplifying event. Currently, the best method for detecting LMP is the galectin puncta formation assay which can be used for a broad range of sample types, both fixed and live, is easy to perform, and highly sensitive. This method, which is similar to the widely used LC3 puncta formation assay for autophagy, is based on the translocation of galectins to damaged lysosomes resulting in a change from uniform to punctate staining pattern. Here, we provide protocols for the galectin puncta formation assay in fixed and live cells and for an alternative assay based on fluorescent dextran release from damaged lysosomes, which can be performed in parallel.

Keywords: Cell death; Dextran; EGFP-galectin-3; Galectin; Immunocytochemistry; Lysosomal membrane permeabilization; Lysosome; Lysosome-associated membrane protein; Microscopy.