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Evaluation of cryo-preserved skin tissues using two-photon microscopy

: Riemann, I.; Beier, A.; Schwarz, M.; Dörr, D.; Stracke, F.; Zimmermann, H.


Kollias, N. ; Society of Photo-Optical Instrumentation Engineers -SPIE-, Bellingham/Wash.:
Photonic therapeutics and diagnostics VI : 23 - 25 January 2010, San Francisco, California, United States; SPIE Photonics West
Bellingham, WA: SPIE, 2010 (Proceedings of SPIE 7548)
ISBN: 978-0-8194-7944-0
ISSN: 1605-7422
Paper 754804
Conference "Photonic Therapeutics and Diagnostics" <6, 2010, San Francisco/Calif.>
Photonics West Conference <2010, San Franciso/Calif.>
Conference Paper
Fraunhofer IBMT ()

If no fresh skin samples can be obtained or used, it is important for research and industries to have models and stored tissue samples as close to the native state as possible at disposal. One way to preserve tissues for a longer timeframe is to use deep freezing cryo-techniques. Unfortunately much damage can be induced during the cooling and the thawing processes like disruption of cells and extra-cellular matrices due to the formation of ice crystals. This could lead to a disturbance of the united cell structure up to the point of a loss of cell viability. Two-photon microscopy is able to gather information about cells and tissue components via excitation of the autofluorescence deep inside the sample with a high resolution in both, frozen and thawed states. It is possible to monitor the samples before and after and, important, observe events during the freezing process like the formation of ice crystals. To determine the state of skin tissues after slow rate freezing and the quick process of vitrification, the samples were examined with two-photon microscopy. To establish an optimized freezing-protocol for skin tissues, morphological changes, changes in autofluorescence of endogenous fluorophores (NADH, keratin, flavins, elastin) or changes in second harmonic generation of collagen fibres could provide information about the quality of the used freezing parameters and protective additives and lead to an optimized freezing-protocol with a new set of parameters to obtain mostly intact tissue samples. Multiphoton microscopy has been established as a useful tool for optical in situ quality control of frozen tissues.