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Journal of Biomedical Optics • Open Access

Calibration and validation of an optical sensor for intracellular oxygen measurements

Paper Abstract

Calibration of fluorescent optical sensors for accurate, quantitative intracellular measurements in vivo suffers from lack of a representative medium that appropriately simulates the molecular complexity of the cytosol. We present a novel protocol for accurate intracellular oxygen sensing via fluorescence lifetime imaging microscopy (FLIM) using cell lysate-FLIM measurements to correct the in vitro calibration of a fluorescent oxygen sensor, and we describe electron paramagnetic resonance (EPR) validation studies. Lysate-FLIM studies provided biochemical information, while EPR provided a "gold standard" for intracellular oxygen estimation. Oxygen levels were evaluated in living human normal squamous and adenocarcinoma esophageal epithelial cells, and good agreement was observed between oxygen levels derived from the optical protocol and EPR. The proposed protocol introduces the concept of a living cell line as a reference for estimating unknown oxygen levels in other cell lines and accounts for high degrees of variability between different cell lines.

Paper Details

Date Published: 1 March 2009
PDF: 3 pages
J. Biomed. Opt. 14(2) 020506 doi: 10.1117/1.3116714
Published in: Journal of Biomedical Optics Volume 14, Issue 2
Show Author Affiliations
Dhruv Sud, Univ. of Michigan (United States)
Mary-Ann Mycek, Univ. of Michigan (United States)

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