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- Guldbrand, Stina, 1970
(författare)
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Non-linear Optical Microscopy and Spectroscopy for Biomedical Studies
- 2012
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- This thesis is based on the application of non-linear optical microscopy and spectroscopy techniques within biomedical research. Non-linear optical microscopy gives the possibility of exciting fluorophores using near infrared light. This is an advantage when working with biological tissue, which has low absorption in this wavelength area, making up an ”open window”for non-invasive three dimensional imaging. Of particular interest has been the study of fluorescent xenobiotics in human skin using two-photon fluorescence laser scanning microscopy. The background is the desire to develop new non-invasive tools to study topical drug delivery and improve the understanding of mechanisms involved in contact allergy. In addition, two-photon fluorescence microscopy is a potential tool for non-invasive skin cancer diagnostics, which also is a topic of this thesis. In order to acquire quantitative data, two-photon fluorescence microscopy has been combined with fluorescence correlation spectroscopy (TPFCS). This is to the best of my knowledge the first time TPFCS has been applied to study the diffusion and distribution of fluorescent molecules in human skin.By the use of this method a reactive compound, acting as a contact allergen, has been demonstrated to bind to proteins in the top epidermal layers of the skin, resulting in a significantly slower diffusion. It has been proposed that endogenously formed protoporphyrin IX (PpIX) can be applied to improve contrast when performing two-photon fluorescence microscopy for diagnostics of non-melanoma skin cancer. In this thesis, it is demonstrated that detection of two-photon excited fluorescence of endogenous PpIX in human skin is not possible. Instead, it is preferable to use a slightly shorter wavelength, i.e. 710 nm, to induce one-photon anti-Stokes fluorescence. This finding is of great importance for continued work in the field, bringing non-linear optical microscopy into the clinics. Plasmonic noble metal nanoparticles, e.g. gold nanoparticles, have been proposed as contrast enhancers for several biomedical applications. In this thesis, gold nanoparticles have been explored with respect to their multiphoton induced luminescence when combined with non-linear optical microscopy. By investigating 10 nm gold nanoparticles deposited on glass plates, it is here demonstrated that aggregation and short inter-particle distances are prerequisites in order to detect multiphoton induced luminescence. Thus detection of single particles in a biological environment is unlikely, and future work should be undertaken to explore how the clustering can be controlled in a biological environment to, e.g, be used as a contrast mechanism.
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| 2. |
- Kirejev, Vladimir, 1984, et al.
(författare)
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Multiphoton microscopy – a powerful tool in skin research and topical drug delivery science
- 2012
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Ingår i: Journal of Drug Delivery Science and Technology. - 1773-2247. ; 22:3, s. 250-259
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Tidskriftsartikel (refereegranskat)abstract
- Multiphoton microscopy (MPM) has become a powerful complementary tool in biomedical research, enabling non-invasive three-dimensional imaging of tissue with high resolution. The major advantage is that investigations and visualization can be performed without mechanical destruction of the sample through tissue sectioning. This review will give a brief introduction to the technology, accompanied by examples of how the technique can be implemented within the field of skin research. Specifically, MPM has already made it possible to visualize cellular morphology and the cutaneous distribution of topically applied compounds applied to intact skin. MPM provides information that can be used to assess the bioavailability of drugs and to visualize drug penetration pathways into skin. MPM has also been implemented as a tool for obtaining non-invasive tissue biopsy based on skin autofluorescence in connection to diagnostics of skin cancer. We will also briefly present some recent results where MPM has been used to track cyclodextrin based drugs applied topically. Finally, we will discuss some future challenges of the technology, including label-free imaging, multimodal techniques, and quantitative imaging.
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