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Meeting Abstract

Affinity labeling of thin sections: still a versatile tool for correlative light and electron microscopy


Schwarz,  H
Electron Microscopy, Max Planck Institute for Developmental Biology, Max Planck Society;

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Schwarz, H., & Humbel, B. (2014). Affinity labeling of thin sections: still a versatile tool for correlative light and electron microscopy. In 18th International Microscopy Conference (pp. 71).

Cite as: https://hdl.handle.net/21.11116/0000-000D-2FF5-B
Cyanobacteria form a very important bacterial phylum that emerged at the dawn of life on Earth. These organisms evolved an efficient photoautotrophic metabolism based on oxidation of water molecules and the reduction of CO2 to organic matter using energy of photons. This unique process involves an endogenous thylakoid membrane system and requires two large chlorophyll-protein assemblies called photosystem I and II (PSI and PSII). In the last decades an amazing amount of progress has been achieved in understanding the structure of photosystems, as well as the mechanisms by which they are assembled from individual components. In contrast, we have very limited knowledge regarding the membrane compartment(s) in which the core photosystem subunits are synthesized and assembled into mature photosynthetic complexes. It appears that synthesis and early stages of photosystem biogenesis are inherently related to biogenesis of the whole thylakoid membrane system, and there is an urgent need to localize these processes in the cell.
The unicellular cyanobacterium Synechocystis PCC 6308 provides a unique combination of molecular genetics and physiological characteristics, and is frequently used as a tool for photosynthetic studies. Recently, a distinct region called PratA-defined membrane (PDM) has been identified in Synechocystis in the vicinity of the plasma membrane, and appears to be a likely site for both commencement of PSII biogenesis, and the terminal steps of chlorophyll biosynthesis. At this time, available data regarding PDM are inconclusive, however in an attempt to clarify this, we have commenced localizing the proteins critical for synthesis of chlorophyll-binding PSII subunits such as In correlative microscopy, light microscopy provides the overview and orientation in the complex cells and tissue, while electron microscopy offers the detailed localization and correlation to subcellular structures. High quality electron microscopical preparation methods provide optimal preservation of the cellular ultrastructure. From such preparations serial thin sections are collected and used for comparative histochemical, immunofluorescence and immunogold staining. In light microscopy histological stains identify the orientation of the sample. Immunofluorescence labeling facilitates to identifying the region of interest, namely, the labeled cells expressing the macromolecule under investigation whereas colloidal gold visualize the label within the cellular architecture at high resolution provided by electron microscopy.