Visualizing NBD-lipid Uptake in Mammalian Cells by Confocal Microscopy
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Visualizing NBD-lipid Uptake in Mammalian Cells by Confocal Microscopy. / Baum, Julia F.; Bredegaard, Lasse; Herrera, Sara Abad; Pomorski, Thomas Günther.
In: Bio-protocol, Vol. 13, No. 13, e4771, 2023.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - Visualizing NBD-lipid Uptake in Mammalian Cells by Confocal Microscopy
AU - Baum, Julia F.
AU - Bredegaard, Lasse
AU - Herrera, Sara Abad
AU - Pomorski, Thomas Günther
N1 - Publisher Copyright: © 2023 The Authors.
PY - 2023
Y1 - 2023
N2 - Eukaryotic cells use a series of membrane transporters to control the movement of lipids across their plasma membrane. Several tools and techniques have been developed to analyze the activity of these transporters in the plasma membrane of mammalian cells. Among them, assays based on fluorescence microscopy in combination with fluorescent lipid probes are particularly suitable, allowing visualization of lipid internalization in living cells. Here, we provide a step-by-step protocol for mammalian cell culture, lipid probe preparation, cell labeling, and confocal imaging to monitor lipid internalization by lipid flippases at the plasma membrane based on lipid probes carrying a fluorophore at a short-chain fatty acid. The protocol allows studying a wide range of mammalian cell lines, to test the impact of gene knockouts on lipid internalization at the plasma membrane and changes in lipid uptake during cell differentiation.
AB - Eukaryotic cells use a series of membrane transporters to control the movement of lipids across their plasma membrane. Several tools and techniques have been developed to analyze the activity of these transporters in the plasma membrane of mammalian cells. Among them, assays based on fluorescence microscopy in combination with fluorescent lipid probes are particularly suitable, allowing visualization of lipid internalization in living cells. Here, we provide a step-by-step protocol for mammalian cell culture, lipid probe preparation, cell labeling, and confocal imaging to monitor lipid internalization by lipid flippases at the plasma membrane based on lipid probes carrying a fluorophore at a short-chain fatty acid. The protocol allows studying a wide range of mammalian cell lines, to test the impact of gene knockouts on lipid internalization at the plasma membrane and changes in lipid uptake during cell differentiation.
KW - Confocal microscopy
KW - Lipid transport
KW - Mammalian cells
KW - NBD-lipid
KW - Plasma membrane
U2 - 10.21769/BioProtoc.4771
DO - 10.21769/BioProtoc.4771
M3 - Journal article
C2 - 37456343
AN - SCOPUS:85164813598
VL - 13
JO - Bio-protocol
JF - Bio-protocol
SN - 2331-8325
IS - 13
M1 - e4771
ER -
ID: 382557690