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Towards real-time breast tumor analysis: visualizing key breast structures using THG/SHG microscopy

23 Sep | By Laura van Huizen
Towards real-time breast tumor analysis: visualizing key breast structures using THG/SHG microscopy
Third (green) and second (red) harmonic generation microscopy images of key breast structures (left) show good agreement with the standard histopathology images (right)
Image source: LaserLab Amsterdam, Vrije Universiteit Amsterdam

For patients with cancer, a fast diagnosis and precise excision of the tumor tissue is important. To improve this, a technique that can assess the tumor tissue with a speed that enables ‘live’ feedback to the surgeon while she/he operates would be very helpful.  A novel imaging technique that has been suggested as promising clinical tool, mainly for cancer diagnosis, is third and second harmonic generation (THG/SHG) microscopy. Researchers of the Vrije Universiteit Amsterdam explored the potential of this technique for the assessment of breast tumor tissue.


For cancer instant pathology it is important to visualize the full morphology of healthy and diseased tissue. Relevant histopathological features include cell and nuclear pleomorphism, nuclear-to-cytoplasmic ratio, growth pattern of the cells and fibrosis. For breast tumor diagnosis, especially the identification of cellular features in the glandular tissue is important, because most breast cancers are derived from the epithelial cells in this tissue. Also, the information on collagen orientation and distribution can be helpful for breast tumor diagnosis.


THG and SHG microscopy can provide this morphological information. Both are nonlinear optical microscopy techniques that do not require labeling of tissue to generate 3D images with sub-cellular resolution and intrinsic depth sectioning within seconds. THG contrast is provided by interfaces and inhomogeneities, and therewith reveals cells, including cell nuclei, while SHG contrast is provided by noncentrosymmetric molecular structures, with as main contrast source collagen.


Studies using SHG/THG microscopy to image breast tissue have shown very promising results, but have been applied on fixated tissue or to visualize the stroma only. The researchers of the Vrije Universiteit Amsterdam now showed that SHG/THG microscopy can identify the key breast components without the need of processing the tissue (no labelling and fixation) within a short time span of a few minutes. Glandular tissue, including cells and cell nuclei, fat tissue, connective tissue, blood vessels and nerves were identified, in good agreement with the gold standard for diagnosis, histopathology.


Currently, a study including imaging breast tumor tissue is in progress, to determine the added value for THG/SHG microscopy for the distinction between normal and cancerous breast tissue. Furthermore, at the Vrije Universiteit Amsterdam in a startup company derived from this research group, Tritos Diagnostics, a transportable, ex-vivo THG/SHG microscope has been developed, which will be soon tested in the operation room, in close collaboration with surgeons and pathologists. The research group is also working on automated image analysis, which will greatly aid to the decision-making using the THG/SHG images. Another goal besides ex-vivo analysis, is in-situ multi-photon imaging using micro-endoscopes to aid a surgeon in determining which tissue to excise. In future, SHG/THG in-situ and ex-vivo analysis may well be valuable additions to the medical imaging instrumentarium of the surgeon.


These findings were published in 2019 in the Journal of Biophotonics.

Link to publication


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