Combining Wide-Field Super-Resolution Microscopy and Electron Tomography

Braet, Filip; Cheng, Delfine; Huynh, Minh; Henriquez, Jeffrey; Shami, Gerald J.; Lampe, Marko

doi:10.1016/b978-0-12-801075-4.00007-0

Methods in Cell Biology

2014

DOI: 10.1016/b978-0-12-801075-4.00007-0

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Combining Wide-Field Super-Resolution Microscopy and Electron Tomography

Filip Braet¹,

Delfine Cheng

Minh Huynh

et al.

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Cited by 3 publications

(1 citation statement)

References 27 publications

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“…Specifically, we will correlate molecular-and cellular dynamics data, along with inter-and intra-cellular signalling events of hepatic colorectal metastases obtained by multimodal CM using cross-integrative analysis. The multimodal imaging approaches utilised in this study include: live animal and live cell imaging [3], microcomputed tomography [4], wide-field and super-resolution nanoscopy [5], correlative fluorescence scanning and transmission electron microscopy [6,7], atomic force probing, and 3-D large-and smallvolume electron tomography [8,9] (Figure 1). Furthermore, we will discuss the careful selection of biomolecular labels for cross-correlative imaging within the same sample of the model organism of interest.…”

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confidence: 99%

Dissecting the Cellular Behaviour of Colorectal Cancer via Multimodal Imaging and Correlative Microscopy

Braet

2017

Microsc Microanal

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These days, the common ground between structural cell biology and molecular biology continues to grow thanks to the biomolecular insights offered by correlative microscopy (CM). CM utilizes the latest advances in contemporary multimodal imaging approaches that are now capable of filling in the missing pieces of the puzzle. CM allows us to interrogate a sample's fine structure from the nano-to micrometre scale in a way that is impossible with any one technique alone [1]. The rapidly expanding CM toolset delivers a strong degree of confidence in the observed structures of interest as insights obtained from one microscopy approach can be directly interrogated by other imaging modalities [2].Here, a versatile multimodal CM workflow encompassing vastly different temporal and spatial resolutions will be presented to reveal novel structural-functional information in colorectal cancer (CRC)-mediated events within hepatic tissue, including liver sinusoidal cells. Specifically, we will correlate molecular-and cellular dynamics data, along with inter-and intra-cellular signalling events of hepatic colorectal metastases obtained by multimodal CM using cross-integrative analysis. The multimodal imaging approaches utilised in this study include: live animal and live cell imaging [3], microcomputed tomography [4], wide-field and super-resolution nanoscopy [5], correlative fluorescence scanning and transmission electron microscopy [6,7], atomic force probing, and 3-D large-and smallvolume electron tomography [8,9] (Figure 1). Furthermore, we will discuss the careful selection of biomolecular labels for cross-correlative imaging within the same sample of the model organism of interest.These multimodal imaging and CM approaches added the following insights: (i) metastasizing CRC cells are preferentially restrained in the periportal sinusoids of liver tissue due to their size; (ii) macrophages, natural killer cells, and endothelial cells work in concert as one immune-surveillance guardian in the removal of the majority of the CRC cells, thereby controlling tumour growth; (iii) ~5% of the CRC cells are able to escape the local hepatic immune system and provide themselves a gateway towards the liver parenchymal tissue by inducing apoptosis and retraction of the endothelium; (iv) the apoptosis regulating molecules Fas/FasL play a key role in the metastatic cascade; (v) the IFNγ/NO pathway has been proven to be a promising molecular target for the development of a new immunotherapeutic strategy in CRC; and, (vi) surviving cancer cells are primarily confined to the Space of Disse and to the Glisson capsule, suggesting that further tumour proliferation and growth are likely to initiate from these extracellular matrix-rich regions of the liver.By drawing on these novel correlative image models, we are finally beginning to understand the mechanisms of CRC metastasis that will enable us to strategically design alternative therapeutic approaches for the treatment of colorectal liver metastases and assess their efficacy [10]. Key approaches ...

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Dissecting the Cellular Behaviour of Colorectal Cancer via Multimodal Imaging and Correlative Microscopy

Braet

2017

Microsc Microanal

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Correlative light and electron microscopy methods for the study of virus–cell interactions

et al. 2016

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Electron microscopy (EM) is an invaluable tool to study the interactions of viruses with cells, and the ultrastructural changes induced in host cells by virus infection. Light microscopy (LM) is a complementary tool with the potential to locate rare events, label specific components, and obtain dynamic information. The combination of LM and EM in correlative light and electron microscopy (CLEM) is particularly powerful. It can be used to complement a static EM image with dynamic data from live imaging, identify the ultrastructure observed in LM, or, conversely, provide molecular specificity data for a known ultrastructure. Here, we describe methods and strategies for CLEM, discuss their advantages and limitations, and review applications of CLEM to study virus-host interactions.Keywords: correlative light and electron microscopy; electron microscopy; virus-host interaction Viruses are obligate intracellular parasites whose replication is dependent on the intimate interaction with the host cell. Virus infection starts upon target cell engagement, generally through the recognition of attachment factors and specific receptor molecules on the cell surface. A combination of intrinsic features, such as virus size and the presence of a lipid envelope, with specific factors, such as the ability to recruit distinct host receptors, determine the type of internalization process. Whatever the entry route is, that is, endocytosis or direct fusion at the plasma membrane, the result is the release of the viral genome, often bound to viral proteins, into the cytoplasm. There, the viral genome must reach the appropriate intracellular Abbreviations CEMOVIS, cryo-electron microscopy of vitreous sections; CLEM, correlative light and electron

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Correlated light and electron microscopy observations of the uterine epithelial cell actin cytoskeleton using fluorescently labeled resin-embedded sections

Moore

Cheng

Shami

et al. 2016

Micron

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Combining Wide-Field Super-Resolution Microscopy and Electron Tomography

Cited by 3 publications

References 27 publications

Dissecting the Cellular Behaviour of Colorectal Cancer via Multimodal Imaging and Correlative Microscopy

Dissecting the Cellular Behaviour of Colorectal Cancer via Multimodal Imaging and Correlative Microscopy

Correlative light and electron microscopy methods for the study of virus–cell interactions

Correlated light and electron microscopy observations of the uterine epithelial cell actin cytoskeleton using fluorescently labeled resin-embedded sections

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