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  1. Article ; Online: cryoWriter: a blotting free cryo-EM preparation system with a climate jet and cover-slip injector.

    Rima, Luca / Zimmermann, Michael / Fränkl, Andri / Clairfeuille, Thomas / Lauer, Matthias / Engel, Andreas / Engel, Hans-Andreas / Braun, Thomas

    Faraday discussions

    2022  Volume 240, Page(s) 55–66

    Abstract: Electron microscopy (EM) introduced a fast and lasting change to structural and cellular biology. However, the sample preparation is still the bottleneck in the cryogenic electron microscopy (cryo-EM) workflow. Classical specimen preparation methods ... ...

    Abstract Electron microscopy (EM) introduced a fast and lasting change to structural and cellular biology. However, the sample preparation is still the bottleneck in the cryogenic electron microscopy (cryo-EM) workflow. Classical specimen preparation methods employ a harsh paper-blotting step, and the protein particles are exposed to a damaging air-water interface. Therefore, improved preparation strategies are urgently needed. Here, we present an amended microfluidic sample preparation method, which entirely avoids paper blotting and allows the passivation of the air-water interface during the preparation process. First, a climate jet excludes oxygen from the sample environment and controls the preparation temperature by varying the relative humidity of the grid environment. Second, the integrated "coverslip injector" allows the modulation of the air-water interface of the thin sample layer with effector molecules. We will briefly discuss the climate jet's effect on the stability and dynamics of the sample thin films. Furthermore, we will address the coverslip injector and demonstrate significant improvement in the sample quality.
    MeSH term(s) Cryoelectron Microscopy ; Proteins ; Specimen Handling ; Microfluidics ; Water
    Chemical Substances Proteins ; Water (059QF0KO0R)
    Language English
    Publishing date 2022-11-08
    Publishing country England
    Document type Journal Article
    ISSN 1364-5498
    ISSN (online) 1364-5498
    DOI 10.1039/d2fd00066k
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article ; Online: Microfluidics and Electron Microscopy

    Luca Rima / Andri Fränkl / Anastasia Syntychaki / Paola Oliva / M. Zimmermann / Xavier Wildermuth / Rosemarie Sütterlin / Thomas Braun

    CHIMIA, Vol 74, Iss

    A Powerful Couple

    2020  Volume 11

    Keywords Cryo-em ; Microfluidics ; Nanoanalytics ; Single particle analysis ; Single-cell analysis ; Visual proteomics ; Chemistry ; QD1-999
    Language German
    Publishing date 2020-11-01T00:00:00Z
    Publisher Swiss Chemical Society
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article: Electron microscopy for ultrastructural analysis and protein localization in

    Frankl, Andri / Mari, Muriel / Reggiori, Fulvio

    Microbial cell (Graz, Austria)

    2015  Volume 2, Issue 11, Page(s) 412–428

    Abstract: ... The ... ...

    Abstract The yeast
    Language English
    Publishing date 2015-10-12
    Publishing country Austria
    Document type Journal Article ; Review
    ZDB-ID 2814756-X
    ISSN 2311-2638
    ISSN 2311-2638
    DOI 10.15698/mic2015.11.237
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  4. Article ; Online: Structural investigation of ACE2 dependent disassembly of the trimeric SARS-CoV-2 Spike glycoprotein

    Ni, Dongchun / Lau, Kelvin / Lehmann, Frank / Fränkl, Andri / Hacker, David / Pojer, Florence / Stahlberg, Henning

    bioRxiv

    Abstract: The human membrane protein Angiotensin-converting enzyme 2 (hACE2) acts as the main receptor for host cells invasion of the new coronavirus SARS-CoV-2. The viral surface glycoprotein Spike binds to hACE2, which triggers virus entry into cells. As of ... ...

    Abstract The human membrane protein Angiotensin-converting enzyme 2 (hACE2) acts as the main receptor for host cells invasion of the new coronavirus SARS-CoV-2. The viral surface glycoprotein Spike binds to hACE2, which triggers virus entry into cells. As of today, the role of hACE2 for virus fusion is not well understood. Blocking the transition of Spike from its prefusion to post-fusion state might be a strategy to prevent or treat COVID-19. Here we report a single particle cryo-electron microscopy analysis of SARS-CoV-2 trimeric Spike in presence of the human ACE2 ectodomain. The binding of purified hACE2 ectodomain to Spike induces the disassembly of the trimeric form of Spike and a structural rearrangement of its S1 domain to form a stable, monomeric complex with hACE2. This observed hACE2 dependent dissociation of the Spike trimer suggests a mechanism for the therapeutic role of recombinant soluble hACE2 for treatment of COVID-19.
    Keywords covid19
    Publisher BioRxiv; WHO
    Document type Article ; Online
    DOI 10.1101/2020.10.12.336016
    Database COVID19

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  5. Article ; Online: Electron microscopy for ultrastructural analysis and protein localization in Saccharomyces cerevisiae

    Andri Frankl / Muriel Mari / Fulvio Reggiori

    Microbial Cell, Vol 2, Iss 11, Pp 412-

    2015  Volume 428

    Abstract: The yeast Saccharomyces cerevisiae is a key model system for studying of a multitude of cellular processes because of its amenability to genetics, molecular biology and biochemical procedures. Ultrastructural examinations of this organism, though, are ... ...

    Abstract The yeast Saccharomyces cerevisiae is a key model system for studying of a multitude of cellular processes because of its amenability to genetics, molecular biology and biochemical procedures. Ultrastructural examinations of this organism, though, are traditionally difficult because of the presence of a thick cell wall and the high density of cytoplasmic proteins. A series of recent methodological and technical developments, however, has revived interest in morphological analyses of yeast (e.g. [1-3]). Here we present a review of established and new methods, from sample preparation to imaging, for the ultrastructural analysis of S. cerevisiae. We include information for the use of different fixation methods, embedding procedures, approaches for contrast enhancement, and sample visualization techniques, with references to successful examples. The goal of this review is to guide researchers that want to investigate a particular process at the ultrastructural level in yeast by aiding in the selection of the most appropriate approach to visualize a specific structure or subcellular compartment.
    Keywords electron microscopy ; electron tomography ; immunolabeling ; chemical fixation ; cryo-immobilization ; correlative light and electron microscopy ; Saccharomyces cerevisiae ; Biology (General) ; QH301-705.5
    Language English
    Publishing date 2015-10-01T00:00:00Z
    Publisher Shared Science Publishers OG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  6. Article ; Online: Maximized song learning of juvenile male zebra finches following BDNF expression in the HVC.

    Dittrich, Falk / Ter Maat, Andries / Jansen, Rene F / Pieneman, Anton / Hertel, Moritz / Frankl-Vilches, Carolina / Gahr, Manfred

    The European journal of neuroscience

    2013  Volume 38, Issue 9, Page(s) 3338–3344

    Abstract: During song learning, vocal patterns are matched to an auditory memory acquired from a tutor, a process involving sensorimotor feedback. Song sensorimotor learning and song production of birds is controlled by a set of interconnected brain nuclei, the ... ...

    Abstract During song learning, vocal patterns are matched to an auditory memory acquired from a tutor, a process involving sensorimotor feedback. Song sensorimotor learning and song production of birds is controlled by a set of interconnected brain nuclei, the song control system. In male zebra finches, the beginning of the sensorimotor phase of song learning parallels an increase of the brain-derived neurotrophic factor (BDNF) in just one part of the song control system, the forebrain nucleus HVC. We report here that transient BDNF-mRNA upregulation in the HVC results in a maximized copying of song syllables. Each treated bird shows motor learning to an extent similar to that of the selected best learners among untreated zebra finches. Because this result was not found following BDNF overexpression in the target areas of HVC within the song system, HVC-anchored mechanisms are limiting sensorimotor vocal learning.
    MeSH term(s) Animals ; Brain-Derived Neurotrophic Factor/genetics ; Brain-Derived Neurotrophic Factor/metabolism ; Finches ; Learning ; Male ; RNA, Messenger/genetics ; RNA, Messenger/metabolism ; Somatosensory Cortex/metabolism ; Somatosensory Cortex/physiology ; Up-Regulation ; Vocalization, Animal
    Chemical Substances Brain-Derived Neurotrophic Factor ; RNA, Messenger
    Language English
    Publishing date 2013-11
    Publishing country France
    Document type Journal Article
    ZDB-ID 645180-9
    ISSN 1460-9568 ; 0953-816X
    ISSN (online) 1460-9568
    ISSN 0953-816X
    DOI 10.1111/ejn.12329
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  7. Article ; Online: Regulatory mechanisms of testosterone-stimulated song in the sensorimotor nucleus HVC of female songbirds.

    Dittrich, Falk / Ramenda, Claudia / Grillitsch, Doris / Frankl-Vilches, Carolina / Ko, Meng-Ching / Hertel, Moritz / Goymann, Wolfgang / ter Maat, Andries / Gahr, Manfred

    BMC neuroscience

    2014  Volume 15, Page(s) 128

    Abstract: Background: In male birds, influence of the sex steroid hormone testosterone and its estrogenic metabolites on seasonal song behavior has been demonstrated for many species. In contrast, female song was only recently recognized to be widespread among ... ...

    Abstract Background: In male birds, influence of the sex steroid hormone testosterone and its estrogenic metabolites on seasonal song behavior has been demonstrated for many species. In contrast, female song was only recently recognized to be widespread among songbird species, and to date, sex hormone effects on singing and brain regions controlling song development and production (song control nuclei) have been studied in females almost exclusively using domesticated canaries (Serinus canaria). However, domesticated female canaries hardly sing at all in normal circumstances and exhibit only very weak, if any, song seasonally under the natural photoperiod. By contrast, adult female European robins (Erithacus rubecula) routinely sing during the winter season, a time when they defend feeding territories and show elevated circulating testosterone levels. We therefore used wild female European robins captured in the fall to examine the effects of testosterone administration on song as well as on the anatomy and the transcriptome of the song control nucleus HVC (sic). The results obtained from female robins were compared to outcomes of a similar experiment done in female domesticated canaries.
    Results: Testosterone treatment induced abundant song in female robins. Examination of HVC transcriptomes and histological analyses of song control nuclei showed testosterone-induced differentiation processes related to neuron growth and spacing, angiogenesis and neuron projection morphogenesis. Similar effects were found in female canaries treated with testosterone. In contrast, the expression of genes related to synaptic transmission was not enhanced in the HVC of testosterone treated female robins but was strongly up-regulated in female canaries. A comparison of the testosterone-stimulated transcriptomes indicated that brain-derived neurotrophic factor (BDNF) likely functions as a common mediator of the testosterone effects in HVC.
    Conclusions: Testosterone-induced singing of female robins correlated with cellular differentiation processes in the HVC that were partially similar to those seen in the HVC of testosterone-treated female canaries. Other modes of testosterone action, notably related to synaptic transmission, appeared to be regulated in a more species-specific manner in the female HVC. Divergent effects of testosterone on the HVC of different species might be related to differences between species in regulatory mechanisms of the singing behavior.
    MeSH term(s) Animals ; Brain/anatomy & histology ; Brain/blood supply ; Brain/physiology ; Brain-Derived Neurotrophic Factor/metabolism ; Female ; Gene Expression Profiling ; Intermediate Filaments/metabolism ; Microarray Analysis ; Neurons/cytology ; Neurons/physiology ; Random Allocation ; Seasons ; Songbirds/physiology ; Testosterone/metabolism ; Vocalization, Animal/physiology
    Chemical Substances Brain-Derived Neurotrophic Factor ; Testosterone (3XMK78S47O)
    Language English
    Publishing date 2014-12-02
    Publishing country England
    Document type Comparative Study ; Journal Article
    ISSN 1471-2202
    ISSN (online) 1471-2202
    DOI 10.1186/s12868-014-0128-0
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