In vivo optical biopsy of hamster oral cavity with epi-third-harmonic-generation microscopy

Abstract

The first in vivo optical virtual biopsy based on epi-third-harmonic-generation (THG) microscopy is successfully demonstrated using Syrian hamster oral mucosa as a model system. Without complex physical biopsy procedures, epi-THG microscopy can provide high spatial resolution dynamic images of oral mucosa and sub-mucosa in all three dimensions. The demonstrated intra-vital epi-THG microscopy provide high resolution observation of blood flow in the capillary and could be a promising tool to image angiogenesis, which is an important feature for many human diseases including malignancies. The system setup of epi-THG microscopy can be easily integrated with other nonlinear optical microscopy such as second-harmonic generation and multi-photon fluorescence microscopy by using the same laser system to provide better integrated molecular and structural information for future clinical diagnosis. By adding 6% acetic acid solution on the mucosa, THG contrast on the borders of nuclei was found to be greatly enhanced due to the alterations of their linear and nonlinear THG susceptibilities. With a virtual-transition-based technology without using fluorescence, the optical epi-THG biopsy we demonstrated shows promise for future noninvasive in vivo diseases examinations.

Third harmonic generation microscopy of cells and tissue organization

“…Exogenous application of acetic acid, which reversibly crosslinks nucleoproteins, can further enhance THG contrast in intact nuclei by changing their refractive index . This can be exploited to improve the detection of ( pre-)cancerous lesions in mucosal tissues with optical biopsies, based on the presence of abnormalities in nuclear morphology and nucleocytoplasmic ratios that are typical for malignant cells Tai et al, 2006;Tsai et al, 2011).…”

“…THG has the potential to complement these existing imaging approaches. The applicability of THG in optical biopsies has been demonstrated in hamster and human oral mucosa, as well as human skin and adipose tissue (Chen et al, 2009;Tai et al, 2005Tai et al, , 2006Tsai et al, 2011Tsai et al, , 2013. Furthermore, the image contrast generated using THG is suitable for discriminating melanoma and basal cell carcinoma lesions from normal skin and benign nevi .…”

Applying tattoo dye as a third-harmonic generation contrast agent forin vivooptical virtual biopsy of human skin

“…Harmonic generation microscopy (HGM) has emerged as an important imaging modality in biological study, [1][2][3][4][5][6][7][8][9] especially in in vivo applications [10][11][12][13][14][15][16][17] because of its nonlinear and noninvasive nature. Our preliminary in vivo clinical studies indicated that HGM can provide a sub-micron spatial resolution, low photodamage and phototoxicity, reduced dye toxicity as a result of the minimized use of external fluorophores, and low photobleaching.…”

“…Our preliminary in vivo clinical studies indicated that HGM can provide a sub-micron spatial resolution, low photodamage and phototoxicity, reduced dye toxicity as a result of the minimized use of external fluorophores, and low photobleaching. In human skin, third-harmonic generation (THG) microscopy can provide intrinsic contrasts in elastic fibers, 17 cytoplasmic membrane, 13 nucleus, 13 actin filaments, 18 lipid bodies, 19 hemoglobin, 20 and melanin. 21 With the combination of THG and second-harmonic generation (SHG) processes, multi-harmonic generation microscopy (HGM) is an ideal imaging tool for morphological visualizations of human skin.…”

Imaging colloidal particle induced topological defects in a nematic liquid crystal using third harmonic generation microscopy

“…Indeed, an interface in either refractive index and/or χ (3) is required for significant THG [12,13]. This property of THG is used in label free imaging in both biology and the material sciences [14][15][16][17][18]. Due to the nonlinear nature of the interaction, THG has inherent optical sectioning properties and thus true three-dimensional imaging capability.…”