Biocenter Oulu: technology services for biomedical research
Biocenter Oulu offers services for the study of proteins, cells and genes, and the generation creation of transgenic animals. One of its strengths is the light and electron microscope imaging of tissues and cells.
According to Professor Lauri Eklund, Coordinator in Light Microscopy at Biocenter Oulu, genetically modified model animals and mice in particular have helped researchers to understand more about phenomena related to the normal development and tissue operation of mammals than of any other organism. In addition to better understanding of developmental processes, these can be used as model organisms to study human diseases.
“Many imaging projects in Oulu concern the study of genetically modified mice. Mouse embryos and organs are imaged in Oulu either in whole, or by means of tissue specimens and light microscopy down to the accuracy level of individual cells, or even in higher resolution using electron microscope. We have also introduced methods to obtain images of cells and macromolecular structures in aestheticized mice, enabling us to see living tissues at high resolution. For this purpose, an intravital imaging laboratory has been set up, which enables cell examination in animals under anaesthesia. Small surgical procedures can also be performed in the laboratory.
“In addition, we use image data to create 3D models by means of optical sectioning. In addition to imaging of relatively large volumes, motorised microscopes can also be used to create tissue images of a large surface areausing tile scanning.”
The core facility service of Biocenter Oulu’s light microscopy specialises in what is known as mesoscopic imaging. Mesoscopic imaging helps to understand interaction between cells in a complex tissue environment, or even in entire organisms. Samples of mesoscopic scale are larger in volume and area than normal microscopy —ranging from a few millimetres to a couple of centimetres. These include mouse embryos, organoids resembling three-dimensional organs, and entire small model organisms, such as flies and fish emryos.
“In technical terms, mesoscopic imaging often requires a tissue culture environment suitable for microscopic imaging, specially designed 3D imaging equipment, tissue clarification methods, and advanced image analysis and processing capacity,” says Eklund.
Locating cellular and tissue structures and events
Biocenter Oulu has a range of microscopes that enable various imaging methods and can pinpoint the location of various events in cells and tissues. A time dimension (4D imaging) can be added to three-dimensional images in living samples. Image sequences can be created, for example to show how cells differentiate and grow into, say, embryos or organoids.
“Thanks to the work done by Professor SeppoVainio’s research group, we can grow an organoid within a few days, using various types of renal progenitor cells. This is something that has attracted international interest. Many researchers have come to Oulu to learn about the technique.”
Confocal and light sheet fluorescence microscopes are suitable for the imaging of three-dimensional and living samples. Especailly light sheet fluorescence microscope can scan the samples quickly without phototocix effect. Electron microscopY, on the other hand, can be used to find changes in the structures of cells and extracellular spaces, which are beyond the resolution of optical microscopy. However, this technology requires that the samples are fixed into place.
Although light waves cannot create magnifications in the same way as electron microscopes, innovative use of excitation laser light, fluorescently labelled molecules and image data processing, can achieve a level of resolution in optical microscopy which allows the examination of individual cells, cell organelles and macromolecular structures.
In most cases such objects must be made fluorescent in order to become visible in 3D microscopy. In the living cells or organism a fluorescent protein can beattached genetically, to the molecule to be studied. The fluorescent compounds (fluorophores) absorb energy from the excitation light and release part of this energy in the form of longer light wavelengths. This quantum mechanics phenomenon, which is visible to the human eye in certain range, is known as fluorescence.
It is also possible to search for specific proteins in cells and tissues by using antibodies to which a fluorescent marker has been attached. The antibody will identify its epitope in a certain protein and attach to it. Once it has attached, the marker can be detected with a microscope. The marker is chosen based on the kind of microscope that will be used to study the sample.
“We have microscopes equipped with spectral detectors and continuous laser light that enable us to study more than one fluorescent label simultaneously. This enables the study of complex interactions and multiple proteins.”
In fluorescence microscopy, fluorescent molecules are used as markers, while electron microscopy uses electron dense material, such as gold, for example.
”Oulu also has what is known as a label free imaging method that does not require additional markers or contrast agents, but can image endogenous molecules These include connective tissue collagen made visible with multiphoton technology, or other body’s own molecules, such as haemoglobin, which can berevealed through photoacoustic imaging. In the case of the latter technology, by combining excitation lasers, structural and functional information can be obtained from tissues, such as the structure of blood vessels and the blood’s oxygenation level.
“These technologies are very useful when imaging living tissues into which it is difficult to introduce markers.”
In terms of electron microscopy, Oulu specialises in the ultrastructural pathology in model organisms and immunoelectron microscopy of tissues and cultured cells,. These techniques provide information on the minutest details and the exact position of the proteins being studied in cell and tissue structures.
In immunoelectron microscopy, a metal labelled antibody or other reagent is related to the protein being studied, which means that the protein’s location can be determined very accurately, in a nanometer scale. This can provide new information on, for example, cell structures and protein interactions or orientation.
“The use of electron microscope methods to examine ultrastructures has been particularly fruitful in terms of the study of macromolecular structures of the extracellular matrix, which cannot be seen using optical microscopy. A fairly new area of study involves extracellular vesicles, “exomes”, which can be imaged by means of electron microscopy.
Data analysis challenges
The problems with traditional imaging have been low resolution, low imaging depth, and lack of effective and quantitative analytics for the image data. In addition, special experience is required in order to extract biological data from the images.
Computer learning and machine vision methodsfor image interpretation has been developed in Oulu. In the approaches Biocenter Oulu has been collaborating with Professor Janne Heikkilä from the University of Oulu’s Center for Machine Vision and Signal Analysis.
“Data storage, transfer and analyses are challenging with respect to the 3D and 4D imaging of large samples. When data is transferred from microscope to user, one should be able to analyse it. Depending on the data, analyses may require a high amount of computing power. If the original data is stored somewhere distant, image processing may be slowed down due to insufficient data transfer speeds.”
Lauri Eklund regards the infrastructure provided by ELIXIR Finland’s CSC as the best solution, in national terms, for raw data storage and the reuse of open data.
Although metadata is linked to image data, there still can be many problems with data management.
“In order for image data to be reusable, it should conform to certain standards and be curated and annotated. Ideally, research infrastructures should provide image data “librarians” and “image information specialists”.
Biocenter Oulu is part of Biocenter Finland, which coordinates infrastructural activities for major national research. It is also a member of various European research infrastructures. These are Infrafrontier (transgenic mice), Euro-BioImaging (biological imaging) and Instruct (protein structure research).
ELIXIR is building an infrastructure to support research and the bio sector. It combines the leading organisations of 21 European countries and the European Molecular Biology Laboratory (EMBL) into a single infrastructure for biological information. Its Finnish centre is CSC – IT Center for Science Ltd. http://www.elixir-finland.org http://www.elixir-europe.org