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Methods Mol Biol. 2019 Feb 16. doi: 10.1007/7651_2019_212. [Epub ahead of print]

Step-by-Step Protocol for Superparamagnetic Nanoparticle-Based Endosome and Lysosome Isolation from Eukaryotic Cell.

Author information

1
EnvirotransgeneĀ® Bio-solutions Global Chennai India and Institute of Cancer Research, Oslo University Hospital, Oslo, Norway. deepak.balaji@bitsaa.org.
2
Laboratory of Lipid Metabolism and Cancer, Louvain, Belgium.
3
Bannari Amman Institute of Technology, Sathyamangalam, TN, India.
4
Faculty of Biology, Hanoi National University of Education, Hanoi, Vietnam.
5
Institute for Research and Development, Duy Tan University, Danang, Vietnam.
6
School of Odonto Stomatology, Hanoi Medical University, Hanoi, Vietnam.
7
School of Bio Sciences and Technology (SBST), VIT University, Vellore, TN, India.

Abstract

Here, we report our step-by-step protocol for superparamagnetic nanoparticle (SPMNP)-based endosome and lysosome isolation from HeLa. Briefly, we synthesized SPMNP 1.0 with iron oxide (Fe3O4) core using thermal decomposition method. Further, we performed ligand-exchange strategy for surface functionalization of SPMNP 1.0 with dimercaptosuccinic acid (DMSA). Thus, we generated DMSA-SPMNP 2.0 and used DMSA-SPMNP 2.0 to isolate endosomes and lysosome from HeLa cells. Using our SPMNP subcellular fractionation protocol, we are able to isolate high-pure-high-yield lysosomes using DMSA-SPMNP 2.0 for lysosome proteomics and lipidomics in order to better understand subcellular compartments.

KEYWORDS:

DMSA; Endosomes; Lysosomes and pulse-chase; Superparamagnetic nanoparticles

PMID:
30771190
DOI:
10.1007/7651_2019_212

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