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Journal of Biomedical Optics • Open Access

Label-free separation of human embryonic stem cells and their differentiating progenies by phasor fluorescence lifetime microscopy
Author(s): Chiara Stringari; Robert Sierra; Peter J. Donovan; Enrico Gratton

Paper Abstract

We develop a label-free optical technique to image and discriminate undifferentiated human embryonic stem cells (hESCs) from their differentiating progenies in vitro. Using intrinsic cellular fluorophores, we perform fluorescence lifetime microscopy (FLIM) and phasor analysis to obtain hESC metabolic signatures. We identify two optical biomarkers to define the differentiation status of hESCs: Nicotinamide adenine dinucleotide (NADH) and lipid droplet-associated granules (LDAGs). These granules have a unique lifetime signature and could be formed by the interaction of reactive oxygen species and unsaturated metabolic precursor that are known to be abundant in hESC. Changes in the relative concentrations of these two intrinsic biomarkers allow for the discrimination of undifferentiated hESCs from differentiating hESCs. During early hESC differentiation we show that NADH concentrations increase, while the concentration of LDAGs decrease. These results are in agreement with a decrease in oxidative phosphorylation rate. Single-cell phasor FLIM signatures reveal an increased heterogeneity in the metabolic states of differentiating H9 and H1 hESC colonies. This technique is a promising noninvasive tool to monitor hESC metabolism during differentiation, which can have applications in high throughput analysis, drug screening, functional metabolomics and induced pluripotent stem cell generation.

Paper Details

Date Published: 20 April 2012
PDF: 12 pages
J. Biomed. Opt. 17(4) 046012 doi: 10.1117/1.JBO.17.4.046012
Published in: Journal of Biomedical Optics Volume 17, Issue 4
Show Author Affiliations
Chiara Stringari, Univ. of California, Irvine (United States)
Robert Sierra, Univ. of California, Irvine (United States)
Peter J. Donovan, Univ. of California, Irvine (United States)
Enrico Gratton, Univ. of California, Irvine (United States)

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