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Basic research with a focus on application

In the Centre of Applied Photonics (CAP) at the University of Constance, interdisciplinary teams of researchers are working on new optical technologies, with a special focus on laser technology. Basic research also takes into account the potential for application. In partnership with Professor Alfred Leitensdorfer, a specialist in the development of fibre lasers, the biologist Dr. Elisa Ferrando-May is focusing on the development of a new laser-scanning confocal microscope.

Non-linear confocal microscopy is a useful tool for investigating biological processes in living tissue. Conventional devices are best suited for thin and transparent samples, explains Dr. Ferrando-May. The representation of different tissue levels enables scientists to reconstruct a three-dimensional picture. However, laser beams are unable to penetrate thick organs to sufficient depths.
Dr. Elisa Ferrando-May at the University of Constance is in charge of setting up the Bio-Imaging Centre. (Photo: Keller-Ulrich)
Although non-linear as well as multiphoton confocal microscopes use laser light that enables thick tissue specimens to be investigated, the devices are very expensive and difficult to operate. The Constance researchers believe that fibre lasers have properties that might be useful for multiphoton microscopy. Instead of a laser crystal, the fibre lasers use Erbium-doted glass fibres as laser medium. This new technological development would only cost about one third that of a conventional microscope as the “price is largely determined by the light source used,” said Dr. Ferrando-May. As the components used come from the telecommunication industry, they are already in an advanced stage of development, giving the Constance researchers access to reliable and cheap standard components.

Optimal adjustment to different dyes

New laser scanning confocal microscope (Photo: Ferrando-May, University of Constance)
The tool developed by the Constance researchers is not only cheaper, but also a lot easier to operate. The instrument is smaller and more stable, and the optics can be adjusted much more easily than those of conventional microscopes. Moreover, the microscope works with short pulses and the light source can be adjusted in the same way as a radio where different wavelengths can be chosen. However, further research is required before the researchers are able to assess the consequences of the new tool for modern imaging. “At present, we are unable to say what we will be able to see,” said Dr. Ferrando-May. However, initial results are very promising. The Constance researchers have been able to show that their system is on a par with instruments already available on the market and it also provides excellent images of thicker samples, such as for example neurone assemblies.
The new laser microscope also opens up further research possibilities as the whole colour range can be displayed and the wavelength adjusted. This adjustment excites different fluorophores with which the living cells are labelled. Chemists have been developing an increasing number of new fluorophores. However, before they can be used effectively, the laser has to be adapted to the requirements of the specific dyes. “Previously, we had to chose the dyes according to the technical possibilities that were available. Now, the new technique enables us to focus on the biological circumstances,” said Dr. Ferrando-May summarising the advantages of the new system.

Identification of repair proteins

The new laser microscopes are not only suitable for pure imaging, but at the same time enable the manipulation of cells or cell constituents. 3-photon absorption can be used to introduce specific DNA damage and visualise the action of different repair proteins. This is achieved by coupling a specific fluorophore to a repair protein and bringing it into the cell. This enables researchers to follow the action of proteins in living cells.

At the moment, Dr. Ferrando-May and her team are looking into whether the adjustment, i.e. increase of the wavelength might lead to higher selectivity in the type of damage. For example, the Constance method enables the creation of specific light-induced damage in skin cells. Depending on the damage, the specific repair processes and their dynamics can be tracked under the microscope, enabling the scientists to clearly differentiate the repair of DNA strand ruptures or photo products. The researchers’ goal is to use the DNA lesions for the identification of tools that carry out the different repairs.

DNA defects and their repair are ongoing processes. It is estimated that there are about 50,000 DNA single-strand ruptures in each body cell per day. Double-strand ruptures are rare, occurring approximately ten times per day per cell.

The different repair pathways are also important for the understanding of genetic skin diseases. The Constance researchers are working with Professor Hanspeter Nägeli from the University of Zurich on Xeroderma pigmentosum, a skin disease characterised by defective DNA repair enzymes, which prevents damaged skin from regenerating once it has been exposed to UV-light. Normal people can repair such UV-induced DNA damage by inserting new bases into the DNA, Xeroderma pigmentosum patients lack this capacity and are often referred to as midnight children or children of the dark, because they can only go outside after nightfall.

Excellent support

Since microscopy can be used in many disciplines, the University of Constance decided to establish a central microscopy facility. The new Bio-Imaging Centre, under the leadership of Dr. Elisa Ferrando-May, will open its doors in August 2008 and will house a broad range of imaging methods and applications at a single site. Ferrando-May also hopes to start up a cooperation with imaging specialists in the computer science department as the data produced by the laser microscope often have to be quantified in order to enable reliable statements to be made. The biologists, physicists and computer scientists at the University of Constance are also hoping that their project will receive support from the German Excellence Initiative.

mek – 29th June 2008
© BIOPRO Baden-Württemberg GmbH
Further information:
Department of Biology
University of Constance
Universitätsstraße 10
P.O. Box X911
78457 Konstanz
Tel.: +49 (0)7531 88-4045
Fax: +49 (0)7531 88-4033
E-mail: elisa.may@uni-konstanz.de
Website address: https://www.gesundheitsindustrie-bw.de/en/article/news/basic-research-with-a-focus-on-application