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HeLa cells, transfected with Cox8A-SiR, highlighting mitochondrial cristae in living cells. To achieve live-cell compatible superresolution, the membrane-permeable dye silicon rhodamine (SiR) is used.
HeLa cells, transfected with Cox8A-SiR, highlighting mitochondrial cristae in living cells. To achieve live-cell compatible superresolution, the membrane-permeable dye silicon rhodamine (SiR) is used.

For light microscopy projects, users receive training on the respective microscopes followed by special imaging techniques. These techniques range from live cell to intravital imaging, second harmonic generation imaging, fluorescence lifetime imaging, different kinds of superresolution imaging, and more. At present, we are improving our superresolution approaches towards better live-cell compatibility (see Fig. 1). In collaboration with the Chen lab, we tested a newly developed mitochondrial dye for superresolution imaging that is now fixable, showing beautiful cristae morphologies of mitochondria (Chen et al. 2024, PNAS).

Electron tomography reconstruction of a mitochondrion from an osteosarcoma cell (collaboration with U. Brandt)
2. Electron tomography reconstruction of a mitochondrion from an osteosarcoma cell (collaboration with U. Brandt)

In electron microscopy, we provide full-service options, including sample preparation and image acquisition, with the option of EM training for dedicated EM users. Within the last two years, we expanded our TEM acquisition towards electron tomography and serial section acquisition to move towards EM volume imaging and 3D reconstruction (see Fig. 2). We further increased our portfolio in different CLEM approaches (Correlative Light and Electron Microscopy). We were able to relocate rare structures that were observed in tissue within the light microscope and to relocate their ultrastructure precisely within the electron microscope. And we combined the superresolution technique STED (STimulted Emission Depletion) with EM, resulting in Super-CLEM (see Fig 3). In EM, we will focus on more automated image acquisition methods in the future.

Super CLEM in HeLa cells using a new, fixalbe mitochondrial probe. (A) STED image of HeLa cells with PKMO FX (A´) Confocal image of mtDNA (SybrGold). (B) Electron tomograph of the same mitochondria using established pre-embedding CLEM method. (B´) Overlay of fluorescent and electron microscope image. Scalebar: 3 µm (Chen et al. 2024, PNAS)
3. Super CLEM in HeLa cells using a new, fixalbe mitochondrial probe. (A) STED image of HeLa cells with PKMO FX (A´) Confocal image of mtDNA (SybrGold). (B) Electron tomograph of the same mitochondria using established pre-embedding CLEM method. (B´) Overlay of fluorescent and electron microscope image. Scalebar: 3 µm (Chen et al. 2024, PNAS)

During the last year, we took the CMMC Imaging Platform under our umbrella and welcomed a new team member to our group, Dr. Mehrnaz Babaki is located at the CMMC to support users in light microscopy experiments and image analysis as well as CECAD members in image analysis. In addition to the already offered support in image data analysis in our group and the training on dedicated analysis software, we will focus more on automated image analysis approaches and deep-learning algorithms.

Dr. Astrid Schauss CECAD

Dr. Astrid Schauss

Head of CECAD / CMMC Imaging Facility

+49 221 478 840 27

aschauss[at]uni-koeln.de

CECAD Cologne / Imaging Facility

CECAD Forschungszentrum
Joseph-Stelzmann-Str. 26

50931 Köln

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