Methods for Probing Lysosomal Membrane Permeabilization.

Abstract

Cell death triggered by lysosomal membrane permeabilization (LMP) is gaining increased interest as target for cancer therapy, but the death pathway also plays an important role in normal physiology (e.g., during involution of the mammary gland). LMP-induced cell death is triggered by release of hydrolases including cysteine cathepsin proteases from the lysosomal lumen into the cytosol. Limited release of proteases to the cytoplasm induces apoptosis or apoptosis-like cell death, whereas massive LMP results in rapid cellular necrosis. Here we introduce three complementary methods for quantifying and visualizing LMP: (i) monitoring LMP by immunocytochemistry, (ii) visualizing LMP by fluorescent dextran release, and (iii) quantification of LMP by activity measurements of lysosomal enzymes in digitonin-extracted cytosol.

• Authors: Marja Jäättelä, Jesper Nylandsted
• Journal: Cold Spring Harbor protocols (Cold Spring Harb Protoc) Vol. 2015 Issue 11 Pg. 975-8 (Nov 02 2015) ISSN: 1559-6095 [Electronic] United States
• PMID: 26527770 (Publication Type: Journal Article)
• Copyright: © 2015 Cold Spring Harbor Laboratory Press.

Chemical References

• Detergents
• Enzymes
• Fluorescent Dyes
• Digitonin

Topics

• Cell Line
• Detergents (metabolism)
• Digitonin (metabolism)
• Enzymes (analysis)
• Fluorescent Dyes (analysis)
• Humans
• Immunohistochemistry (methods)
• Lysosomes (drug effects, enzymology)
• Membranes (drug effects, physiology)
• Permeability (drug effects)