Open AccessRapid Lipid Droplet Isolation Protocol Using a Well-established Organelle Isolation Kit
Author(s) -
Jascha Brettschneider,
Jason Correnti,
Chun-Yao Lin,
Bianca Williams,
Amanke Oranu,
Amy Kuriakose,
Dru McIver-Jenkins,
Abigail Haba,
Isabelle Kaneza,
Sookyoung Jeon,
Eleonora Scorletti,
Rotonya M. Carr
Publication year - 2019
Publication title -
journal of visualized experiments
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.596
H-Index - 91
ISSN - 1940-087X
DOI - 10.3791/59290
Subject(s) - endoplasmic reticulum , organelle , cytosol , lipid droplet , reagent , microbiology and biotechnology , biochemistry , isolation (microbiology) , biology , chemistry , bioinformatics , enzyme
Lipid droplets (LDs) are bioactive organelles found within the cytosol of the most eukaryotic and some prokaryotic cells. LDs are composed of neutral lipids encased by a monolayer of phospholipids and proteins. Hepatic LD lipids, such as ceramides, and proteins are implicated in several diseases that cause hepatic steatosis. Although previous methods have been established for LD isolation, they require a time-consuming preparation of reagents and are not designed for the isolation of multiple subcellular compartments. We sought to establish a new protocol to enable the isolation of LDs, endoplasmic reticulum (ER), and lysosomes from a single mouse liver. Further, all reagents used in the protocol presented here are commercially available and require minimal reagent preparation without sacrificing LD purity. Here we present data comparing this new protocol to a standard sucrose gradient protocol, demonstrating comparable purity, morphology, and yield. Additionally, we can isolate ER and lysosomes using the same sample, providing detailed insight into the formation and intracellular flux of lipids and their associated proteins.