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Metabolite Profiling and Stable Isotope Tracing in Sorted Subpopulations of Mammalian Cells
journal contribution
posted on 2016-02-08, 00:00 authored by Irena Roci, Hector Gallart-Ayala, Angelika Schmidt, Jeramie Watrous, Mohit Jain, Craig
E. Wheelock, Roland NilssonBiological samples such as tissues,
blood, or tumors are often
complex and harbor heterogeneous populations of cells. Separating
out specific cell types or subpopulations from such complex mixtures
to study their metabolic phenotypes is challenging because experimental
procedures for separation may disturb the metabolic state of cells.
To address this issue, we developed a method for analysis of cell
subpopulations using stable isotope tracing and fluorescence-activated
cell sorting followed by liquid chromatography–high-resolution
mass spectrometry. To ensure a faithful representation of cellular
metabolism after cell sorting, we benchmarked sorted extraction against
direct extraction. While peak areas differed markedly with lower signal
for amino acids but higher signal for nucleotides, mass isotopomer
distributions from sorted cells were generally in good agreement with
those obtained from direct extractions, indicating that they reflect
the true metabolic state of cells prior to sorting. In proof-of-principle
studies, our method revealed metabolic phenotypes specific to T cell
subtypes, and also metabolic features of cells in the committed phase
of the cell division cycle. Our approach enables studies of a wide
range of adherent and suspension cell subpopulations, which we anticipate
will be of broad importance in cell biology and biomedicine.