Detection and Isolation of Apoptotic Bodies to High Purity

Journal of Visualized Experiments : JoVE
Thanh Kha PhanGeorgia K Atkin-Smith

Abstract

Apoptotic bodies (ApoBDs), microvesicles and exosomes are the key members of the extracellular vesicle family, with ApoBDs being one of the largest type. It has been proposed that ApoBDs can aid cell clearance as well as intercellular communication through trafficking biomolecules. Conventional approaches used for the identification and isolation of ApoBDs are often limited by the lack of accurate quantification and low sample purity. Here, we describe a workflow to confirm the induction of apoptosis, validate ApoBD formation, and isolate ApoBDs to high purity. We will also outline and compare fluorescence-activated cell sorting (FACS) and differential centrifugation based approaches to isolate ApoBDs. Furthermore, the purity of isolated ApoBDs will be confirmed using a previously establish flow cytometry-based staining and analytical method. Taken together, using the described approach, THP-1 monocyte apoptosis and apoptotic cell disassembly was induced and validated, and ApoBD generated from THP-1 monocytes were isolated to a purity of 97-99%.

Citations

Sep 2, 2020·Biochemical Society Transactions·Thanh Kha PhanIvan Ka Ho Poon
May 10, 2020·Communications Biology·Georgia K Atkin-SmithIvan K H Poon
Oct 1, 2020·Stem Cell Research & Therapy·Golara NasiriLobat Tayebi
Nov 3, 2020·Frontiers in Cell and Developmental Biology·Maojiao LiWeidong Tian

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