Two‐photon microscopy of deep intravital tissues and its merits in clinical research

Abstract

SummaryMultiphoton excitation laser scanning microscopy, relying on the simultaneous absorption of two or more photons by a molecule, is one of the most exciting recent developments in biomedical imaging. Thanks to its superior imaging capability of deeper tissue penetration and efficient light detection, this system becomes more and more an inspiring tool for intravital bulk tissue imaging. Two-photon excitation microscopy including 2-photon fluorescence and second harmonic generated signal microscopy is the most common multiphoton microscopic application. In the present review we take diverse ocular tissues as intravital samples to demonstrate the advantages of this approach. Experiments with registration of intracellular 2-photon fluorescence and extracellular collagen second harmonic generated signal microscopy in native ocular tissues are focused. Data show that the in-tandem combination of 2-photon fluorescence and second harmonic generated signal microscopy as two-modality microscopy allows for in situ co-localization imaging of various microstructural components in the whole-mount deep intravital tissues. New applications and recent developments of this high technology in clinical studies such as 2-photon-controlled drug release, in vivo drug screening and administration in skin and kidney, as well as its uses in tumourous tissues such as melanoma and glioma, in diseased lung, brain and heart are additionally reviewed. Intrinsic emission two-modal 2-photon microscopy/tomography, acting as an efficient and sensitive non-injurious imaging approach featured by high contrast and subcellular spatial resolution, has been proved to be a promising tool for intravital deep tissue imaging and clinical studies. Given the level of its performance, we believe Correspondence to B.-G. Wang. Tel: +49 3641 938496; fax: +49 3641 938552; e-mail: Baogui.Wang@mti.uni-jena.de that the non-linear optical imaging technique has tremendous potentials to find more applications in biomedical fundamental and clinical research in the near future.

Proximal Tubules Have the Capacity to Regulate Uptake of Albumin

“…2,12,14,22 One of the main functions of the PTC is retrieval of filtered proteins and our studies, and other recent reports, support the importance of the PTC in regulating albumin excretion rate. 2 of TR-RSA uptake in all surface PTs of control (n=3 rats, 157 fields quantified) and protein overloaded (n=8 rats, 176 fields quantified) rats.…”

Imaging Strategies for Tissue Engineering Applications

“…Reprinted with permission from Bago et al 104 Color images available online at www.liebertpub.com/teb constructs are enabled with the penetration depth increased up to *500 mm. 59,82 For instance, two-photon fluorescence microscopy has been successfully applied to visualize the structure of polymer gels and engineered tissue at different depths, even in a bioreactor system (Fig. 7A).…”

In vivo imaging of cell nuclei by photoacoustic microscopy without staining

“…The high image contrast results from a difference between strong light absorption of nucleic acids and weak absorption of protein, which reaches the maximum at a wavelength of 250 nm. In addition to UV-PAM, several other modern optical microscopy technologies have been explored for in vivo imaging of unstained cell nuclei, such as reflectance confocal microscopy [8][9][10][11], multiphoton microscopy [12][13][14], and third-harmonic generation microscopy [15,16], However, reflectance confocal microscopy is difficult to provide specific image contrast for nuclei [8], multiphoton microscopy produces in vivo images of cell nuclei with negative contrast [13], and third harmonic generation microscopy generates nuclear images with low contrast [15].…”