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Intravital imaging of gastrointestinal diseases in preclinical models using two-photon laser scanning microscopy

Abstract

Two-photon laser scanning microscopy (TPLSM), relying on two-photon excitation restricted to the focal plane, has become the gold standard in biomedical research because of its ability to produce high-resolution, higher penetrating imaging of biological materials over an extended duration without an significant photobleaching. Intravital (in vivo) imaging using TPLSM for intra-abdominal organs has long been a technical challenge because of the difficulty of achieving high-quality and higher magnification imaging with acceptable cardiac and respiratory motion artifacts. A method of intravital TPLSM was developed using an organ-stabilizing system for imaging the intra-abdominal organs of green fluorescent protein transgenic mice. This method was further refined for the time-series imaging of intra-abdominal organs in the same mouse model using intravital TPLSM. These procedures allow the observation of not only a single cell or tissue microenvironment at a higher penetrating depth for a longer period of time but also to observe the same organ of the same mouse at multiple time points in preclinical models. This report presents the general principles and properties of TPLSM for biomedical research. In addition, the methods and the usefulness of time-series intravital TPLSM imaging for preclinical gastrointestinal diseases is discussed.

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Acknowledgments

This work was supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan (KAKENHI 22591484 to K. Tanaka, 21591723 to Y. Inoue, and 21390377 to M. Kusunoki). This work was also supported by a grant-in-aid of The Public Trust Fund for the Promotion of Surgery, Tokyo, Japan. K. Tanaka was supported by the Okasan-Kato Foundation with regard to this work, and was also supported by the Ichiro Kanehara Foundation.

Conflict of interest

The authors have no conflict of interest to disclose.

Author information

Correspondence to Koji Tanaka.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary Movie 3b: z-stacks imaging of normal liver bar = 50 μm. TPLSM = two-photon laser scanning microscopy. (WMV 990 kb)

Supplementary Movie 3a: time-lapse imaging of normal liver (WMV 1326 kb)

Supplementary Movie 3b: z-stacks imaging of normal liver bar = 50 μm. TPLSM = two-photon laser scanning microscopy. (WMV 990 kb)

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Tanaka, K., Toiyama, Y., Inoue, Y. et al. Intravital imaging of gastrointestinal diseases in preclinical models using two-photon laser scanning microscopy. Surg Today 43, 123–129 (2013) doi:10.1007/s00595-012-0283-9

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Keywords

  • Gastrointestinal disease
  • Intravital imaging
  • Two-photon laser scanning microscopy
  • Preclinical model