Advertisement

Respiration Rate Determined by Phosphorescence-Based Sensors

  • Michael KonopkaEmail author
Protocol
Part of the Springer Protocols Handbooks book series (SPH)

Abstract

Respiration rates can be a powerful diagnostic tool that provides insight into the metabolic activity of the cells. An optical-based method is well suited for making oxygen consumption measurements in microbial populations, whether on a model organism or environmental sample. This approach utilizes phosphorescent dyes since the lifetime of their excited state depends on the oxygen concentration. Two systems are described using closed sample chambers which can be constructed at minimal costs from off-the-shelf parts. The first system is designed around a glass cuvette utilizing an oil layer as an oxygen barrier. The second system is adapted to an existing microscope and uses a cavity well slide with a glass coverslip lid as the sample chamber. In both systems a photomultiplier tube or gated CCD camera is used for detecting the phosphorescent signal which, when calibrated, provides a reliable measurement of oxygen concentration over time.

Keywords:

Environmental microbiology Oxygen sensor Phosphorescence lifetime Respiration rate measurement 

References

  1. 1.
    Otterstedt K, Larsson C, Bill RM et al (2004) Switching the mode of metabolism in the yeast Saccharomyces cerevisiae. EMBO Rep 5:532–537CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Ramamoorthy R, Dutta PK, Akbar SA (2003) Oxygen sensors: materials, methods, designs, and applications. J Mater Sci 38:4271–4282CrossRefGoogle Scholar
  3. 3.
    Strovas TJ, Dragavon JM, Hankins TJ, Callis JB, Burgess LW, Lidstrom ME (2006) Measurement of respiration rates of Methylobacterium extorquens AM1 cultures by use of a phosphorescence-based sensor. Appl Environ Microbiol 72:1692–1695CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Arain S, John GT, Krause C, Gerlach J, Wolfbeis OS, Klimant I (2006) Characterization of microtiterplates with integrated optical sensors for oxygen and pH, and their applications to enzyme activity screening, respirometry, and toxicological assays. Sens Actuators B 113:639–648CrossRefGoogle Scholar
  5. 5.
    Will Y, Hynes J, Ogurtsov VI, Papkovsky DB (2006) Analysis of mitochondrial function using phosphorescent oxygen-sensitive probes. Nat Protoc 1:2563–2572CrossRefPubMedGoogle Scholar
  6. 6.
    Lopes AS, Greve T, Callesen H (2007) Quantification of embryo quality by respirometry. Theriogenology 67:21–31CrossRefPubMedGoogle Scholar
  7. 7.
    Wilson DF, Vanderkooi JM, Green TJ, Maniara G, DeFeo SP, Bloomgarden DC (1987) A versatile and sensitive method for measuring oxygen. Adv Exp Med Biol 215:71–77CrossRefPubMedGoogle Scholar
  8. 8.
    Han BH, Manners I, Winnik MA (2005) Phosphorescence quenching of dyes adsorbed to silica thin-layer chromatography plates. Anal Chem 77:8075–8085CrossRefPubMedGoogle Scholar
  9. 9.
    O’Riordan TC, Buckley D, Ogurtsov V, O’Connor R, Papkovsky DB (2000) A cell viability assay based on monitoring respiration by optical oxygen sensing. Anal Biochem 278:221–227CrossRefPubMedGoogle Scholar
  10. 10.
    Molter TW, Holl MR, Dragavon JM et al (2008) A new approach for measuring single-cell oxygen consumption rates. IEEE Trans Autom Sci Eng 5:32–42CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Strovas TJ, McQuaide SC, Anderson JB et al (2010) Direct measurement of oxygen consumption rates from attached and unattached cells in a reversibly sealed, diffusionally isolated sample chamber. Adv Biosci Biotechnol 5:398–408CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Chan SP, Fuller ZJ, Demas JN, DeGraff BA (2001) Optimized gating scheme for rapid lifetime determinations of single-exponential luminescence lifetimes. Anal Chem 73:4486–4490CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  1. 1.Department of ChemistryUniversity of AkronAkronUSA

Personalised recommendations