Display Settings:

Format

Send to:

Choose Destination
Photochem Photobiol. 2004 Jul-Aug;80:1-6.

Spatial control of reactive oxygen species formation in fibroblasts using two-photon excitation.

Author information

  • 1Department of Dermatology, University of California-San Francisco, San Francisco, CA, USA.

Abstract

Two-photon excitation (2PE) provides a means of generating reactive oxygen species (ROS) in cells and tissues with a high degree of spatial specificity. In cultured monolayers of human fibroblasts and fibroblast-derived cells treated with the commonly used probe of ROS formation, 5-(and 6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate, acetyl ester (CM-H(2)DCFDA), cells irradiated through a microscope objective with 150 fs near-infrared laser pulses became highly fluorescent, reflecting intracellular ROS formation. The fluorescence intensity inside cells increased quadratically with the average power of radiation for pulsed excitation and was unchanged for continuous wave irradiation with the same average power. Single fibroblasts embedded within dermal equivalents were also targeted in this manner and formed ROS, whereas neighboring unirradiated cells were spared. These results demonstrate that ROS can be generated intracellularly in skin cells using 2PE of the metabolic or oxidative products of CM-H(2)DCFDA and that formation of ROS can be localized in both cell monolayers and in a tissue equivalent. This technique should be useful in understanding the response of whole tissues such as skin to local generation of ROS and may have applications in photodynamic therapy.

PMID:
15339206
[PubMed - indexed for MEDLINE]
PMCID:
PMC2774523
Free PMC Article

Images from this publication.See all images (5)Free text

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Icon for Blackwell Publishing Icon for PubMed Central
    Loading ...
    Write to the Help Desk