Triplet Imaging of Oxygen Consumption during the Contraction of a Single Smooth Muscle Cell (A7r5)

QC 20110207
The measurement of tissue and cell oxygenation is important for understanding cell metabolism. We have addressed this problem with a novel optical technique, called triplet imaging, that exploits oxygen-induced triplet lifetime changes and is compatible with a variety of fluorophores. A modulated excitation of varying pulse widths allows the extraction of the lifetime of the essentially dark triplet state using a high-fluorescence signal intensity. This enables the monitoring of fast kinetics of oxygen concentration in living cells combined with high temporal and spatial resolution. First, the oxygen-dependent triplet-state quenching of tetramethylrhodamine is validated and then calibrated in an L-ascorbic acid titration experiment demonstrating the linear relation between triplet lifetime and oxygen concentration according to the Stern-Volmer equation. Second, the method is applied to a biological cell system, employing as reporter a cytosolic fusion protein of beta-galactosidase with SNAP-tag labeled with tetramethylrhodamine. Oxygen consumption in single smooth muscle cells A7r5 during an [Arg(8)]-vasopressin-induced contraction is measured. The results indicate a consumption leading to an intracellular oxygen concentration that decays monoexponentially with time. The proposed method has the potential to become a new tool for investigating oxygen metabolism at the single cell and the subcellular level.

Spielmann, ThiemoKTH,Experimentell biomolekylär fysik(Swepub:kth)u176i2oc (författare)
Formey, Aurlie (författare)
Maerki, Iwan (författare)
Leutenegger, Marcel (författare)
Hinz, Boris (författare)
Johnsson, Kai (författare)
Van De Ville, Dimitri (författare)
Lasser, Theo (författare)
KTHExperimentell biomolekylär fysik (creator_code:org_t)

Av författaren/redakt...: Geissbuehler, Ma ...; Spielmann, Thiem ...; Formey, Aurlie; Maerki, Iwan; Leutenegger, Mar ...; Hinz, Boris; visa fler...; Johnsson, Kai; Van De Ville, Di ...; Lasser, Theo; visa färre...

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