@article{6029,
  abstract     = {Protein micropatterning has become an important tool for many biomedical applications as well as in academic research. Current techniques that allow to reduce the feature size of patterns below 1 μm are, however, often costly and require sophisticated equipment. We present here a straightforward and convenient method to generate highly condensed nanopatterns of proteins without the need for clean room facilities or expensive equipment. Our approach is based on nanocontact printing and allows for the fabrication of protein patterns with feature sizes of 80 nm and periodicities down to 140 nm. This was made possible by the use of the material X-poly(dimethylsiloxane) (X-PDMS) in a two-layer stamp layout for protein printing. In a proof of principle, different proteins at various scales were printed and the pattern quality was evaluated by atomic force microscopy (AFM) and super-resolution fluorescence microscopy.},
  author       = {Lindner, Marco and Tresztenyak, Aliz and Fülöp, Gergö and Jahr, Wiebke and Prinz, Adrian and Prinz, Iris and Danzl, Johann G and Schütz, Gerhard J. and Sevcsik, Eva},
  issn         = {22962646},
  journal      = {Frontiers in Chemistry},
  publisher    = {Frontiers Media S.A.},
  title        = {{A fast and simple contact printing approach to generate 2D protein nanopatterns}},
  doi          = {10.3389/fchem.2018.00655},
  volume       = {6},
  year         = {2019},
}

@article{6052,
  abstract     = {Expansion microscopy is a relatively new approach to super-resolution imaging that uses expandable hydrogels to isotropically increase the physical distance between fluorophores in biological samples such as cell cultures or tissue slices. The classic gel recipe results in an expansion factor of ~4×, with a resolution of 60–80 nm. We have recently developed X10 microscopy, which uses a gel that achieves an expansion factor of ~10×, with a resolution of ~25 nm. Here, we provide a step-by-step protocol for X10 expansion microscopy. A typical experiment consists of seven sequential stages: (i) immunostaining, (ii) anchoring, (iii) polymerization, (iv) homogenization, (v) expansion, (vi) imaging, and (vii) validation. The protocol presented here includes recommendations for optimization, pitfalls and their solutions, and detailed guidelines that should increase reproducibility. Although our protocol focuses on X10 expansion microscopy, we detail which of these suggestions are also applicable to classic fourfold expansion microscopy. We exemplify our protocol using primary hippocampal neurons from rats, but our approach can be used with other primary cells or cultured cell lines of interest. This protocol will enable any researcher with basic experience in immunostainings and access to an epifluorescence microscope to perform super-resolution microscopy with X10. The procedure takes 3 d and requires ~5 h of actively handling the sample for labeling and expansion, and another ~3 h for imaging and analysis.},
  author       = {Truckenbrodt, Sven M and Sommer, Christoph M and Rizzoli, Silvio O and Danzl, Johann G},
  journal      = {Nature Protocols},
  number       = {3},
  pages        = {832–863},
  publisher    = {Nature Publishing Group},
  title        = {{A practical guide to optimization in X10 expansion microscopy}},
  doi          = {10.1038/s41596-018-0117-3},
  volume       = {14},
  year         = {2019},
}

@article{8618,
  abstract     = {The reversibly switchable fluorescent proteins (RSFPs) commonly used for RESOLFT nanoscopy have been developed from fluorescent proteins of the GFP superfamily. These proteins are bright, but exhibit several drawbacks such as relatively large size, oxygen-dependence, sensitivity to low pH, and limited switching speed. Therefore, RSFPs from other origins with improved properties need to be explored. Here, we report the development of two RSFPs based on the LOV domain of the photoreceptor protein YtvA from Bacillus subtilis. LOV domains obtain their fluorescence by association with the abundant cellular cofactor flavin mononucleotide (FMN). Under illumination with blue and ultraviolet light, they undergo a photocycle, making these proteins inherently photoswitchable. Our first improved variant, rsLOV1, can be used for RESOLFT imaging, whereas rsLOV2 proved useful for STED nanoscopy of living cells with a resolution of down to 50 nm. In addition to their smaller size compared to GFP-related proteins (17 kDa instead of 27 kDa) and their usability at low pH, rsLOV1 and rsLOV2 exhibit faster switching kinetics, switching on and off 3 times faster than rsEGFP2, the fastest-switching RSFP reported to date. Therefore, LOV-domain-based RSFPs have potential for applications where the switching speed of GFP-based proteins is limiting.},
  author       = {Gregor, Carola and Sidenstein, Sven C. and Andresen, Martin and Sahl, Steffen J. and Danzl, Johann G and Hell, Stefan W.},
  issn         = {2045-2322},
  journal      = {Scientific Reports},
  keywords     = {Multidisciplinary},
  publisher    = {Springer Nature},
  title        = {{Novel reversibly switchable fluorescent proteins for RESOLFT and STED nanoscopy engineered from the bacterial photoreceptor YtvA}},
  doi          = {10.1038/s41598-018-19947-1},
  volume       = {8},
  year         = {2018},
}

@article{5998,
  abstract     = {Genome amplification and cellular senescence are commonly associated with pathological processes. While physiological roles for polyploidization and senescence have been described in mouse development, controversy exists over their significance in humans. Here, we describe tetraploidization and senescence as phenomena of normal human placenta development. During pregnancy, placental extravillous trophoblasts (EVTs) invade the pregnant endometrium, termed decidua, to establish an adapted microenvironment required for the developing embryo. This process is critically dependent on continuous cell proliferation and differentiation, which is thought to follow the classical model of cell cycle arrest prior to terminal differentiation. Strikingly, flow cytometry and DNAseq revealed that EVT formation is accompanied with a genome-wide polyploidization, independent of mitotic cycles. DNA replication in these cells was analysed by a fluorescent cell-cycle indicator reporter system, cell cycle marker expression and EdU incorporation. Upon invasion into the decidua, EVTs widely lose their replicative potential and enter a senescent state characterized by high senescence-associated (SA) β-galactosidase activity, induction of a SA secretory phenotype as well as typical metabolic alterations. Furthermore, we show that the shift from endocycle-dependent genome amplification to growth arrest is disturbed in androgenic complete hydatidiform moles (CHM), a hyperplastic pregnancy disorder associated with increased risk of developing choriocarinoma. Senescence is decreased in CHM-EVTs, accompanied by exacerbated endoreduplication and hyperploidy. We propose induction of cellular senescence as a ploidy-limiting mechanism during normal human placentation and unravel a link between excessive polyploidization and reduced senescence in CHM.},
  author       = {Velicky, Philipp and Meinhardt, Gudrun and Plessl, Kerstin and Vondra, Sigrid and Weiss, Tamara and Haslinger, Peter and Lendl, Thomas and Aumayr, Karin and Mairhofer, Mario and Zhu, Xiaowei and Schütz, Birgit and Hannibal, Roberta L. and Lindau, Robert and Weil, Beatrix and Ernerudh, Jan and Neesen, Jürgen and Egger, Gerda and Mikula, Mario and Röhrl, Clemens and Urban, Alexander E. and Baker, Julie and Knöfler, Martin and Pollheimer, Jürgen},
  issn         = {1553-7404},
  journal      = {PLOS Genetics},
  number       = {10},
  publisher    = {Public Library of Science},
  title        = {{Genome amplification and cellular senescence are hallmarks of human placenta development}},
  doi          = {10.1371/journal.pgen.1007698},
  volume       = {14},
  year         = {2018},
}

@article{6499,
  abstract     = {Expansion microscopy is a recently introduced imaging technique that achieves super‐resolution through physically expanding the specimen by ~4×, after embedding into a swellable gel. The resolution attained is, correspondingly, approximately fourfold better than the diffraction limit, or ~70 nm. This is a major improvement over conventional microscopy, but still lags behind modern STED or STORM setups, whose resolution can reach 20–30 nm. We addressed this issue here by introducing an improved gel recipe that enables an expansion factor of ~10× in each dimension, which corresponds to an expansion of the sample volume by more than 1,000‐fold. Our protocol, which we termed X10 microscopy, achieves a resolution of 25–30 nm on conventional epifluorescence microscopes. X10 provides multi‐color images similar or even superior to those produced with more challenging methods, such as STED, STORM, and iterative expansion microscopy (iExM). X10 is therefore the cheapest and easiest option for high‐quality super‐resolution imaging currently available. X10 should be usable in any laboratory, irrespective of the machinery owned or of the technical knowledge.},
  author       = {Truckenbrodt, Sven M and Maidorn, Manuel and Crzan, Dagmar and Wildhagen, Hanna and Kabatas, Selda and Rizzoli, Silvio O},
  issn         = {1469-3178},
  journal      = {EMBO reports},
  number       = {9},
  publisher    = {EMBO},
  title        = {{X10 expansion microscopy enables 25‐nm resolution on conventional microscopes}},
  doi          = {10.15252/embr.201845836},
  volume       = {19},
  year         = {2018},
}

@article{145,
  abstract     = {Aged proteins can become hazardous to cellular function, by accumulating molecular damage. This implies that cells should preferentially rely on newly produced ones. We tested this hypothesis in cultured hippocampal neurons, focusing on synaptic transmission. We found that newly synthesized vesicle proteins were incorporated in the actively recycling pool of vesicles responsible for all neurotransmitter release during physiological activity. We observed this for the calcium sensor Synaptotagmin 1, for the neurotransmitter transporter VGAT, and for the fusion protein VAMP2 (Synaptobrevin 2). Metabolic labeling of proteins and visualization by secondary ion mass spectrometry enabled us to query the entire protein makeup of the actively recycling vesicles, which we found to be younger than that of non-recycling vesicles. The young vesicle proteins remained in use for up to ~ 24 h, during which they participated in recycling a few hundred times. They were afterward reluctant to release and were degraded after an additional ~ 24–48 h. We suggest that the recycling pool of synaptic vesicles relies on newly synthesized proteins, while the inactive reserve pool contains older proteins.},
  author       = {Truckenbrodt, Sven M and Viplav, Abhiyan and Jähne, Sebsatian and Vogts, Angela and Denker, Annette and Wildhagen, Hanna and Fornasiero, Eugenio and Rizzoli, Silvio},
  issn         = {0261-4189},
  journal      = {The EMBO Journal},
  number       = {15},
  publisher    = {Wiley},
  title        = {{Newly produced synaptic vesicle proteins are preferentially used in synaptic transmission}},
  doi          = {10.15252/embj.201798044},
  volume       = {37},
  year         = {2018},
}

@article{159,
  abstract     = {L-type Ca2+ channels (LTCCs) play a crucial role in excitation-contraction coupling and release of hormones from secretory cells. They are targets of antihypertensive and antiarrhythmic drugs such as diltiazem. Here, we present a photoswitchable diltiazem, FHU-779, which can be used to reversibly block endogenous LTCCs by light. FHU-779 is as potent as diltiazem and can be used to place pancreatic β-cell function and cardiac activity under optical control.},
  author       = {Fehrentz, Timm and Huber, Florian and Hartrampf, Nina and Bruegmann, Tobias and Frank, James and Fine, Nicholas and Malan, Daniela and Danzl, Johann G and Tikhonov, Denis and Sumser, Maritn and Sasse, Philipp and Hodson, David and Zhorov, Boris and Klocker, Nikolaj and Trauner, Dirk},
  journal      = {Nature Chemical Biology},
  number       = {8},
  pages        = {764 -- 767},
  publisher    = {Nature Publishing Group},
  title        = {{Optical control of L-type Ca2+ channels using a diltiazem photoswitch}},
  doi          = {10.1038/s41589-018-0090-8},
  volume       = {14},
  year         = {2018},
}

@article{9229,
  author       = {Danzl, Johann G},
  issn         = {2500-2295},
  journal      = {Opera Medica et Physiologica},
  number       = {S1},
  pages        = {11},
  publisher    = {Lobachevsky State University of Nizhny Novgorod},
  title        = {{Diffraction-unlimited optical imaging for synaptic physiology}},
  doi          = {10.20388/omp2018.00s1.001},
  volume       = {4},
  year         = {2018},
}