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Cited by 207Cancer, Hypoxia, and MetabolismOptical Imaging and Spectroscopy TechniquesAnalytical Chemistry and Sensors

The DOI matches its Crossref record; 77 of the 78 references it cites were found in a registry or an index.

[1] Measurement of absolute oxygen levels in cells and tissues using oxygen sensors and 2-nitroimidazole EF5

Koch, Cameron J.·2002·Methods in Enzymology
DOI 10.1016/s0076-6879(02)52003-6
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Abstract

We have established basic methods, using quantitative measures of EF5 binding, to estimate the actual pO2 of cells and tissues. In situations where the tissue can be dissociated into single cells, or for cell cultures, we can measure the distribution of cellular binding rates using flow cytometry and these can be compared with cells treated under pO2S controlled by the spinner vial or thin-film methods in vitro. The flow cytometer is calibrated by staining V79 cells treated with EF5 under "standard" conditions. For intact tissues treated with EF5 in vivo, we need to correct for possible variations in drug exposure (AUC). Frozen sections are stained for EF5 binding and are analyzed by a sensitive (cooled) CCD camera with linear output vs fluorescence [figure: see text] input. The camera has very consistent sensitivity, but the entire optical system, including the camera, can be calibrated by an absolute fluorescence standard (dye in hemocytometer). This system can also be used to measure the fluorescence of the flow cytometer standards, providing a direct link between the two assays. We can measure the maximum binding rate using the tissue cube method, but need to assume an "average" oxygen dependence of binding for intact tissues. The best-fit approximation for existing data is an inverse relationship between binding and pO2, with binding decreasing 50-fold between 0.1 and 10% oxygen. Using these methods, we routinely estimate the minimum pO2 (maximum binding) in experimental rodent and human tumors. In normal tissue models, an excellent correlation is found between near-maximal binding (severe hypoxia) and apoptosis (heart infarct and ductus arteriosus). Some normal tissues (e.g., skeletal muscle) are refractory to both cellular disaggregation and cube calibration methods. To extend the tissue imaging measurements to a complete two- or three-dimensional analysis of the distribution of tissue pO2s requires a substantial additional investment of imaging methods, which are currently being implemented.

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Research works printed but not linked: 16

  1. Brizel, 1996, Cancer Res, 56, 941
  2. Reynolds, 1990, Cancer Res, 56, 5754
  3. Young, 1988, 85, 9533
  4. Clark, 1956, Trans. Am. Soc. Artif. Int. Organs, 2, 41
  5. Chapman, 1983, Cancer Res, 43, 1523
  6. Taylor, 1978, Cancer Res, 38, 2745
  7. Varghese, 1976, Cancer Res, 36, 3761
  8. Franko, 1987, Cancer Res, 47, 5367
  9. Evans, 1996, Cancer Res, 56, 405
  10. Lord, 1993, Cancer Res, 53, 5271
  11. Laughlin, 1996, J. Pharmacol. Exp. Ther, 277, 1049
  12. Evans, 2000, Cancer Res, 60, 2018
  13. Rasey, 1999, J. Nucl. Med, 40, 1072
  14. Busch, 2000, Cancer Res, 60, 2636
  15. Wiebe, 1996, Q. J. Nucl. Med, 40, 270
  16. Evans, 1999, J. Nucl. Med, 41, 327

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Metadata

Authors
Koch, C. J.
Affiliations
University of Pennsylvania (US)
Corresponding author
Koch, Cameron J.
Venue
Methods in Enzymology
Publisher
Elsevier
Published
2002
Volume
352
Pages
3-31
ISSN
0076-6879 (ISSN-L)
DOI
10.1016/s0076-6879(02)52003-6
OpenAlex
W1554643012
Cited by
207
References
78
Citations by year
2026: 4 · 2025: 5 · 2024: 3 · 2023: 8 · 2022: 6 · 2021: 5 · 2020: 7 · 2019: 8 · 2018: 8 · 2017: 4 · 2016: 13 · 2015: 10 · 2014: 10 · 2013: 6 · 2012: 7
FWCI
25.02
Type
book chapter
Language
en
Open access
closed
Locations
  • Methods in enzymology on CD-ROM/Methods in enzymology (publishedVersion)
  • PubMed (publishedVersion)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Cancer, Hypoxia, and Metabolism
  • Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Optical Imaging and Spectroscopy Techniques
  • Physical Sciences › Chemistry › Analytical Chemistry › Analytical Chemistry and Sensors
Keywords
EF5, 2-nitroimidazole, tissue oxygenation, oxygen sensors, hypoxia, apoptosis, flow cytometry, tissue dissociation, fluorescence imaging, tumor oxygenation, V79 cells, ductus arteriosus, skeletal muscle
MeSH terms
Animals, Hypoxia, Biochemistry, Flow Cytometry, Fluorocarbons, Humans, Hydrocarbons, Fluorinated, Immunohistochemistry, Neoplasms, Oxygen, Pressure, Time Factors, Etanidazole, Mice

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