Abstract:
:This protocol measures externalization of aminophospholipids (APLs) to the outside of the plasma membrane using mass spectrometry (MS). APL externalization occurs in numerous events, and it is relevant for transplant medicine, immunity and cancer. In this protocol, externalized APLs are chemically modified by using a cell-impermeable reagent (sulfo-NHS-biotin), and then they are isolated via a liquid:liquid extraction and quantified by reverse-phase liquid chromatography tandem MS (LC-MS/MS) against in-house-generated standards. This protocol describes a complementary method to existing assays that are not quantitative (e.g., annexin V flow cytometry), and it is applicable to the study of membrane reorganization in all cell types during apoptosis (e.g., during development, cancer, psychiatric disorders and other conditions, aging, vesiculation and cell division). The protocol takes ∼2-4 d, including the generation of standards.
journal_name
Nat Protocjournal_title
Nature protocolsauthors
Thomas CP,Clark SR,Hammond VJ,Aldrovandi M,Collins PW,O'Donnell VBdoi
10.1038/nprot.2013.163subject
Has Abstractpub_date
2014-01-01 00:00:00pages
51-63issue
1eissn
1754-2189issn
1750-2799pii
nprot.2013.163journal_volume
9pub_type
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