Oxygen Microscopy by Two-Photon-Excited Phosphorescence

• Published in: Chemphyschem 2008-08, Vol.9 (12), p.1673-1679
• Main Authors: Finikova, Olga S., Lebedev, Artem Y., Aprelev, Alexey, Troxler, Thomas, Gao, Feng, Garnacho, Carmen, Muro, Silvia, Hochstrasser, Robin M., Vinogradov, Sergei A.
• Format: Article
• Language: English
• Subjects: Atomic and molecular collision processes and interactions; Atomic and molecular physics; Biological and medical sciences; Cells, Cultured; dendrimer; Endothelial Cells; energy transfer; Exact sciences and technology; Fundamental and applied biological sciences. Psychology; Fundamental areas of phenomenology (including applications); General aspects, investigation technics, apparatus; Humans; Intramolecular energy transfer; intramolecular dynamics; dynamics of van der waals molecules; Luminescent Measurements; Microscopy - methods; Molecular Structure; Nanostructures; Nonlinear optics; Optical susceptibility, hyperpolarizability; Optics; oxygen; Oxygen - chemistry; phosphorescence; Photochemistry; Photons; Physics; porphyrin; Tissues, organs and organisms biophysics
• ISSN: 1439-4235; 1439-7641
• DOI: 10.1002/cphc.200800296

High‐resolution images of oxygen distributions in microheterogeneous samples are obtained by two‐photon laser scanning microscopy (2P LSM), using a newly developed dendritic nanoprobe with internally enhanced two‐photon absorption (2PA) cross‐section. In this probe, energy is harvested by a 2PA antenna, which passes excitation onto a phosphorescent metalloporphyrin via intramolecular energy transfer. The 2P LSM allows sectioning of oxygen gradients with near diffraction‐limited resolution, and lifetime‐based acquisition eliminates dependence on the local probe concentration. The technique is validated on objects with a priori known oxygen distributions and applied to imaging of pO2 in cells. Oxygen distributions are imaged by two‐photon laser scanning microscopy (2P LSM) using a newly developed two‐photon‐enhanced phosphorescent nanoprobe (see figure). 2P LSM allows visualization of oxygen gradients in 3D with near diffraction‐limited resolution, and lifetime‐based measurements eliminate dependence on the local probe concentration.