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J Biomed Opt. 2018 Oct;23(10):1-14. doi: 10.1117/1.JBO.23.10.107001.

Intracellular oxygen mapping using a myoglobin-mCherry probe with fluorescence lifetime imaging.

Author information

1
National Heart, Lung, and Blood Institute, National Institutes of Health, Laboratory of Advanced Mic, United States.
2
National Heart, Lung, and Blood Institute, National Institutes of Health, Laboratory of Obesity and, United States.
3
Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes, United States.

Abstract

Oxygen (O2) is one of the most important biometabolites. In abundance, it serves as the limiting terminus of aerobic respiratory chains in the mitochondria of higher organisms; in deficit, it is a potent determinant of development and regulation of other physiological and therapeutic processes. Most knowledge on intracellular and interstitial concentration ([O2]) is derived from mitochondria isolated from cells or tissue biopsies, providing detailed but nonnative insight into respiratory chain function. The possible loss of essential metabolites during isolation and disruption of the normal interactions of the organelle with the cytoskeleton may cause these data to misrepresent intact cells. Several optical methodologies were also developed, but they are often unable to detect heterogeneity of metabolic characteristics among different individual cells in the same culture, and most cannot detect heterogeneous consumption within different areas of a single cell. Here, we propose a noninvasive and highly sensitive fluorescence lifetime microscopy probe, myoglobin-mCherry, appropriate to intracellular targeting. Using our probe, we monitor mitochondrial contributions to O2 consumption in A549 nonsmall cell lung cancer cells and we reveal heterogeneous [O2] within the intracellular environments. The mitochondrial [O2] at a single-cell level is also mapped by adding a peptide to target the probe to the mitochondria.

KEYWORDS:

Myo-mCherry; fluorescence lifetime microscopy; intracellular oxygenation; mitochondria; mtMyo-mCherry

PMID:
30298706
PMCID:
PMC6210794
[Available on 2019-10-08]
DOI:
10.1117/1.JBO.23.10.107001
[Indexed for MEDLINE]
Free PMC Article

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