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Simultaneous In Situ Quantification of Two Cellular Lipid Pools Using Orthogonal Fluorescent Sensors
Author(s) -
Liu ShuLin,
Sheng Ren,
O'Connor Matthew J.,
Cui Yang,
Yoon Youngdae,
Kurilova Svetlana,
Lee Daesung,
Cho Wonhwa
Publication year - 2014
Publication title -
angewandte chemie international edition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.831
H-Index - 550
eISSN - 1521-3773
pISSN - 1433-7851
DOI - 10.1002/anie.201408153
Subject(s) - fluorescence , in situ , chemistry , biophysics , membrane , live cell imaging , lipid accumulation , cell , microbiology and biotechnology , biochemistry , biology , physics , organic chemistry , quantum mechanics
Lipids regulate a wide range of biological activities. Since their local concentrations are tightly controlled in a spatiotemporally specific manner, the simultaneous quantification of multiple lipids is essential for elucidation of the complex mechanisms of biological regulation. Here, we report a new method for the simultaneous in situ quantification of two lipid pools in mammalian cells using orthogonal fluorescent sensors. The sensors were prepared by incorporating two environmentally sensitive fluorophores with minimal spectral overlap separately into engineered lipid‐binding proteins. Dual ratiometric analysis of imaging data allowed accurate, spatiotemporally resolved quantification of two different lipids on the same leaflet of the plasma membrane or a single lipid on two opposite leaflets of the plasma membrane of live mammalian cells. This new imaging technology should serve as a powerful tool for systems‐level investigation of lipid‐mediated cell signaling and regulation.