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Quantitative assays for esterified oxylipins generated by immune cells

Morgan, Alwena Haf, Hammond, Victoria Jayne, Morgan, Lloyd, Thomas, Christopher P. ORCID:, Tallman, Keri A., Garcia-Diaz, Yoel Ruslan, McGuigan, Christopher ORCID:, Serpi, Michaela ORCID:, Porter, Ned A., Murphy, Robert C. and O'Donnell, Valerie Bridget ORCID: 2010. Quantitative assays for esterified oxylipins generated by immune cells. Nature Protocols 5 (12) , pp. 1919-1931. 10.1038/nprot.2010.162

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Phospholipid-esterified oxylipins include newly described families of bioactive lipids generated by lipoxygenases in immune cells. Until now, assays for their quantitation were not well developed or widely available. Here, we describe a mass spectrometric protocol that enables accurate measurement of several esterified oxylipins—in particular hydro(pero)xyeicosatetraenoic acids, hydroxyoctadecadienoic acids, hydroxydocosahexaenoic acids and keto-eicosatetraenoic acids—attached to either phosphatidylethanolamine or phosphatidylcholine. Lipids are isolated from cells or tissue using a liquid-phase organic extraction, then analyzed by HPLC–tandem mass spectrometry (LC/MS/MS) in multiple reaction–monitoring mode. The protocol can simultaneously monitor up to 23 species. Generation of standards takes ~2 d. Following this, extraction of 30 samples takes ~3 h, with LC/MS/MS run time of 50 min per sample.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Medicine
Systems Immunity Research Institute (SIURI)
Subjects: Q Science > QR Microbiology > QR180 Immunology
R Medicine > RS Pharmacy and materia medica
Uncontrolled Keywords: Biochemistry and protein analysis; isolation, purification and seperation; spectroscopy and structural analysis; synthetic chemistry
Publisher: Nature Publishing Group
ISSN: 1754-2189
Last Modified: 17 Oct 2022 10:08

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