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Advanced Light and Fluorescence Microscopy

  • State-of-the-art light and fluorescence microscopes
  • Focus on live cell and correlative light and electron cryo-microscopy

About

Email the ALFM Facility

The Advanced Light and Fluorescence Microscopy (ALFM) Facility operates high-end light microscopes and image analysis resources and develops novel technologies for light microscopy. The ALFM Facility is part of the Technology Platform Light Microscopy (TPLM) of Universität Hamburg (UHH). The ALFM Facility develops in close collaboration with the CSSB cryo-EM facility workflows for correlative light and electron cryo-microscopy.

 

Mission

The mission of the ALFM Facility is to support researchers as efficiently as possible for light microscopy and correlative light and electron (cryo-)microscopy projects. To fulfill this mission, the ALFM has the following tasks:

  1. Equipment: To provide access to and to maintain its light microscopy and image analysis equipment and to upgrade the equipment to remain state-of-the-art.
  2. Training and support: To support researchers with choosing the best imaging and image analysis modality for their studies, to train researchers to use the equipment of the ALFM, to support researchers to resolve problems that come up during imaging projects, and to collaboratively carry out imaging projects together with research groups.
  3. Method development:  To carry out projects to develop novel imaging methods that are of use for CSSB.

 

Support

According to the need of the researchers, we offer different levels of support:

  • Independent use of instruments after training
  • Accompanying support during imaging projects
  • Collaborative development and implementation of imaging projects

In addition, we support researchers with image processing and image analysis.

Upon request, users of the facility can also access the laboratory space near the microscopes for sample preparation and mammalian cell culture, which is equipped with:

  • lab work benches
  • safety cabinets
  • CO2 incubators

 

Equipment

All microscopes at the ALFM Facility are in a BSL2 lab.

- Leica SP8 confocal microscope equipped with FALCON FLIM unit
- Leica cryoCLEM widefield microscope
- Leica cryoCLEM Stellaris 8 confocal microscope
- Leica DMi8 inverse live cell widefield microscope equipped with Primo Alveole micropatterning unit
- Leica DMi8 inverse live cell widefield microscope equipped with THUNDER unit
- Leica DM6 B upright widefield microscope #1
- Leica DM6 B upright widefield microscope #2
- Leica DM6 B upgright widefield microscope #3
- Leica SR GSD 3D super-resolution microscope
- Nikon Ti2 spinning disc and SIM microscope equipped with FRAP unit
- Nikon Ti2 spinning disc microscope equipped with SoRa unit and TIRF unit
- Zeiss LSM880 Airyscan confocal microscope
- Zeiss Elyra P1 super-resolution microscope
- Zeiss Lattice Light Sheet microscope
- BSL2 Cell culture hood and incubator for preparing samples for live cell imaging
- Leica M125C stereo microscope
- Analysis computer

 

Team

5

Facility Head

Roland Thünauer

E-Mail

Staff Scientist, Research Engineer

Tim Laugks

E-Mail

Technical Assistant

Larissa Schatz

E-Mail

Scientific Facility Director

Kay Grünewald

E-Mail

Scientific Facility Director

Tim Gilberger

E-Mail

Access

How to start?
For new users, a meeting will be arranged to discuss how the ALFM Facility can support you and how to access equipment at ALFM.
Please contact the ALFM Facility by email (alfm@cssb-hamburg.de) or contact the facility head directly (roland.thuenauer@cssb-hamburg.de) to arrange a meeting or to get more information.

Booking of equipment
Equipment at ALFM Facility is bookable via a PPMS booking calendar: https://ppms.eu/cssb-hamburg/login/?pf=2
Access to the booking calendar will be granted after initial training and signing of the usage conditions: ALFM Usage Rules (PDF, 272KB).

Address
CSSB Centre for Structural Systems Biology
c/o Deutsches Elektronen-Synchrotron DESY
Notkestraße 85, Building 15
D-22607 Hamburg

Facility Operators:
Universität Hamburg (UHH), Leibniz Institute of Virology (LIV), and Bernhard Nocht Institute for Tropical Medicine (BNITM)

Research

The ALFM Facility is open to collaborate with researchers on scientific projects with a particular focus on method development for light microscopy.

Current projects:

CUI: AIM project cryoLM
In this project we develop novel excitation schemes for light cryo-microscopy (cryoLM).
Collaborators: Kay Grünewald, Franz Kärtner, Kartik Ayyer, Rainer Kaufmann and Felix Lehmkühler

Split luciferases for monitoring protein cell surface arrival
In this project we make split luciferases usable for quantifying the cell surface arrival of membrane proteins in live cells in real time.
Collaborators: Wilfried Weber, Winfried Römer

Publications

How to acknowledge the ALFM Facility in publications:

We kindly ask our users to acknowledge the technology platform in publications containing data that were acquired or analyzed at the ALFM Facility. The text below can be used as template:

We thank the Advanced Light and Fluorescence Microscopy (ALFM) Facility at the Centre for Structural Systems Biology (CSSB), in particular XXX, for support with light microscopy image acquisition and analysis.

Publications with ALFM contribution

2023

Yuan B, Scholz J, Wald J, Thuenauer R, Hennell James R, Ellenberg I, Windhorst S, Faix J, Marlovits TC (2023) Structural basis for subversion of host cell actin cytoskeleton during Salmonella infection.  Sci Adv. 9(49):eadj5777. doi: 10.1126/sciadv.adj5777.

Pietsch E, Niedermüller K, Andrews M, Meyer BS, Lenz TL, Wilson DW, Gilberger TW, Burda PC (2023) Disruption of a Plasmodium falciparum patatin-like phospholipase delays male gametocyte exflagellation. Mol Microbiol. doi:10.1111/mmi.15211.

Alio I, Moll R, Hoffmann T, Mamat U, Schaible UE, Pappenfort K, Alawi M, Schie M, Thünauer R, Stamm J, Rohde H, Streit WR (2023) Stenotrophomonas maltophilia affects the gene expression profiles of the major pathogens Pseudomonas aeruginosa and Staphylococcus aureus in an in vitro multispecies biofilm model.
Microbiol Spectr. 11(6):e0085923. doi: 10.1128/spectrum.00859-23.

Pietsch E, Ramaprasad A, Bielfeld S, Wohlfarter Y, Maco B, Niedermüller K, Wilcke L, Kloehn J, Keller MA, Soldati-Favre D, Blackman MJ, Gilberger TW, Burda PC (2023) A patatin-like phospholipase is important for mitochondrial function in malaria parasites. mBio. e0171823. doi: 10.1128/mbio.01718-23.

Sterrenberg VT, Stalling D, Knaack JIH, Soh TK, Bosse JB, Meier C (2023) A TriPPPro-Nucleotide Reporter with Optimized Cell-Permeable Dyes for Metabolic Labeling of Cellular and Viral DNA in Living Cells. Angew Chem Int Ed Engl. 62(38):e202308271. doi: 10.1002/anie.202308271.

Peiseler M, Araujo David B, Zindel J, Surewaard BGJ, Lee WY, Heymann F, Nusse Y, Castanheira FVS, Shim R, Guillot A, Bruneau A, Atif J, Perciani C, Ohland C, Ganguli Mukherjee P, Niehrs A, Thuenauer R, Altfeld M, Amrein M, Liu Z, Gordon PMK, McCoy K, Deniset J, MacParland S, Ginhoux F, Tacke F, Kubes P (2023) Kupffer cell-like syncytia replenish resident macrophage function in the fibrotic liver.
Science. 381(6662):eabq5202. doi: 10.1126/science.abq5202. 

Fischer AAM, Schatz L, Baaske J, Römer W, Weber W, Thuenauer R (2023) Real-time monitoring of cell surface protein arrival with split luciferases.Traffic. 24(10):453-462. doi: 10.1111/tra.12908. 

Burda PC, Ramaprasad A, Bielfeld S, Pietsch E, Woitalla A, Söhnchen C, Singh MN, Strauss J, Sait A, Collinson LM, Schwudke D, Blackman MJ, Gilberger TW (2023) Global analysis of putative phospholipases in Plasmodium falciparum reveals an essential role of the phosphoinositide-specific phospholipase C in parasite maturation. mBio. 14(4):e0141323. doi: 10.1128/mbio.01413-23.

Sassmannshausen J, Bennink S, Distler U, Küchenhoff J, Minns AM, Lindner SE, Burda PC, Tenzer S, Gilberger TW, Pradel G (2023) Comparative proteomics of vesicles essential for the egress of Plasmodium falciparum gametocytes from red blood cells. Mol Microbiol. doi: 10.1111/mmi.15125.

Soh TK, Pfefferle S, Wurr S, von Possel R, Oestereich L, Rieger T, Uetrecht C, Rosenthal M, Bosse JB (2023) A validated protocol to UV-inactivate SARS-CoV-2 and herpesvirus-infected cells. PLoS One. 18(5):e0274065. doi: 10.1371/journal.pone.0274065

Reichelt J, Sachs W, Frömbling S, Fehlert J, Studencka-Turski M, Betz A, Loreth D, Blume L, Witt S, Pohl S, Brand J, Czesla M, Knop J, Florea BI, Zielinski S, Sachs M, Hoxha E, Hermans-Borgmeyer I, Zahner G, Wiech T, Krüger E, Meyer-Schwesinger C (2023) Non-functional ubiquitin C-terminal hydrolase L1 drives podocyte injury through impairing proteasomes in autoimmune glomerulonephritis. Nat Commun. 14(1):2114. doi: 10.1038/s41467-023-37836-8.

Jordan-Paiz A, Martrus G, Steinert FL, Kaufmann M, Sagebiel AF, Schreurs RRCE, Rechtien A, Baumdick ME, Jung JM, Möller KJ, Wegner L, Grüttner C, Richert L, Thünauer R, Schroeder-Schwarz J, van Goudoever JB, Geijtenbeek TBH, Altfeld M, Pals ST, Perez D, Klarenbeek PL, Tomuschat C, Sauter G, Königs I, Schumacher U, Friese MA, Melling N, Reinshagen K, Bunders MJn (2023) CXCR5+PD-1++ CD4+ T cells colonize infant intestines early in life and promote B cell maturation. Cell Mol Immunol. 20(2):201-213. doi: 10.1038/s41423-022-00944-4.

Kaltenbach L, Martzloff P, Bambach SK, Aizarani N, Mihlan M, Gavrilov A, Glaser KM, Stecher M, Thünauer R, Thiriot A, Heger K, Kierdorf K, Wienert S, von Andrian UH, Schmidt-Supprian M, Nerlov C, Klauschen F, Roers A, Bajénoff M, Grün D, Lämmermann T (2023) Slow integrin-dependent migration organizes networks of tissue-resident mast cells. Nat Immunol. 24(6):915-924. doi: 10.1038/s41590-023-01493-2.

Wichers-Misterek JS, Binder AM, Mesén-Ramírez P, Dorner LP, Safavi S, Fuchs G, Lenz TL, Bachmann A, Wilson D, Frischknecht F, Gilberger TW (2023) A Microtubule-Associated Protein Is Essential for Malaria Parasite Transmission. mBio. 14(1):e0331822. doi: 10.1128/mbio.03318-22.

Kohantorabi M, Wagstaffe M, Creutzburg M, Ugolotti A, Kulkarni S, Jeromin A, Krekeler T, Feuerherd M, Herrmann A, Ebert G, Protzer U, Guédez G, Löw C, Thuenauer R, Schlueter C, Gloskovskii A, Keller TF, Di Valentin C, Stierle A, Noei H (2023) Adsorption and Inactivation of SARS-CoV-2 on the Surface of Anatase TiO2(101). ACS Appl Mater Interfaces.  doi: 10.1021/acsami.2c22078.

Wang M, Thuenauer R, Schubert R, Gevorgyan S, Lorenzen K, Brognaro H, Betzel C (2023) Formation kinetics and physicochemical properties of mesoscopic Alpha-Synuclein assemblies modulated by sodium chloride and a distinct pulsed electric field. Soft Matter. doi: 10.1039/d2sm01615j.

Xu M, Antonova M, Salavei P, Illek K, Meléndez AV, Omidvar R, Thuenauer R, Makshakova O, Römer W (2023) Dimeric Lectin Chimeras as Novel Candidates for Gb3-Mediated Transcytotic Drug Delivery through Cellular Barriers. Pharmaceutics. 15(1):225. doi: 10.3390/pharmaceutics15010225.

2022

Franken LE, Rosch R, Laugks U, Grünewald K (2022) Protocol for live-cell fluorescence-guided cryoFIB-milling and electron cryo-tomography of virus-infected cells. STAR Protoc. 3(4):101696. doi: 10.1016/j.xpro.2022.101696.

Bona A, Seifert M, Thünauer R, Zodel K, Frew IJ, Römer W, Walz G, Yakulov TA (2022) MARVEL domain containing CMTM4 affects CXCR4 trafficking. Mol Biol Cell. 33(13):ar116. doi: 10.1091/mbc.E22-05-0152.

Alder A, Wilcke L, Pietsch E, von Thien H, Pazicky S, Löw C, Mesen-Ramirez P, Bachmann A, Burda PC, Kunick C, Sondermann H, Wilson D, Gilberger TW (2023) Functional inactivation of Plasmodium falciparum glycogen synthase kinase GSK3 modulates erythrocyte invasion and blocks gametocyte maturation. J Biol Chem. 298(9):102360. doi: 10.1016/j.jbc.2022.102360.

Vázquez CA, Widerspick L, Thuenauer R, Schneider C, Reimer R, Neira P, Olal C, Heung M, Niemetz L, Lawrence P, Kucinskaite-Kodze I, Redecke L, Escudero-Pérez B (2022) Nipah Virus Infection Generates Ordered Structures in Cellulo. Viruses. 14(7):1523. doi: 10.3390/v14071523.

Thuenauer R, Kühn K, Guo Y, Kotsis F, Xu M, Trefzer A, Altmann S, Wehrum S, Heshmatpour N, Faust B, Landi A, Diedrich B, Dengjel J, Kuehn EW, Imberty A, Römer W (2022) The Lectin LecB Induces Patches with Basolateral Characteristics at the Apical Membrane to Promote Pseudomonas aeruginosa Host Cell Invasion. mBio. 13(3):e0081922. doi: 10.1128/mbio.00819-22.

Caragliano E, Bonazza S, Frascaroli G, Tang J, Soh TK, Grünewald K, Bosse JB, Brune W (2022) Human cytomegalovirus forms phase-separated compartments at viral genomes to facilitate viral replication. Cell Rep. 38(10):110469. doi: 10.1016/j.celrep.2022.110469.

Flomm FJ, Soh TK, Schneider C, Wedemann L, Britt HM, Thalassinos K, Pfitzner S, Reimer R, Grünewald K, Bosse JB (2022) Intermittent bulk release of human cytomegalovirus. PLoS Pathog. 18(8):e1010575. doi: 10.1371/journal.ppat.1010575.

Alder A, Struck NS, Xu M, Johnson JW, Wang W, Pallant D, Cook MA, Rambow J, Lemcke S, Gilberger TW, Wright GD (2022) A non-reactive natural product precursor of the duocarmycin family has potent and selective antimalarial activity. Cell Chem Biol. 29(5):840-853.e6. doi: 10.1016/j.chembiol.2021.10.005.

2021

Wichers JS, Wunderlich J, Heincke D, Pazicky S, Strauss J, Schmitt M, Kimmel J, Wilcke L, Scharf S, von Thien H, Burda PC, Spielmann T, Löw C, Filarsky M, Bachmann A, Gilberger TW (2021) Identification of novel inner membrane complex and apical annuli proteins of the malaria parasite Plasmodium falciparum. Cell Microbiol. 23(9):e13341. doi: 10.1111/cmi.13341.

Lizarrondo J, Klebl DP, Niebling S, Abella M, Schroer MA, Mertens HDT, Veith K, Thuenauer R, Svergun DI, Skruzny M, Sobott F, Muench SP, Garcia-Alai MM (2021) Structure of the endocytic adaptor complex reveals the basis for efficient membrane anchoring during clathrin-mediated endocytosis. Nat Commun. 12(1):2889. doi: 10.1038/s41467-021-23151-7.

Nelson G, Boehm U, Bagley S, Bajcsy P, Bischof J, Brown CM, Dauphin A, Dobbie IM, Eriksson JE, Faklaris O, Fernandez-Rodriguez J, Ferrand A, Gelman L, Gheisari A, Hartmann H, Kukat C, Laude A, Mitkovski M, Munck S, North AJ, Rasse TM, Resch-Genger U, Schuetz LC, Seitz A, Strambio-De-Castillia C, Swedlow JR, Alexopoulos I, Aumayr K, Avilov S, Bakker GJ, Bammann RR, Bassi A, Beckert H, Beer S, Belyaev Y, Bierwagen J, Birngruber KA, Bosch M, Breitlow J, Cameron LA, Chalfoun J, Chambers JJ, Chen CL, Conde-Sousa E, Corbett AD, Cordelieres FP, Nery ED, Dietzel R, Eismann F, Fazeli E, Felscher A, Fried H, Gaudreault N, Goh WI, Guilbert T, Hadleigh R, Hemmerich P, Holst GA, Itano MS, Jaffe CB, Jambor HK, Jarvis SC, Keppler A, Kirchenbuechler D, Kirchner M, Kobayashi N, Krens G, Kunis S, Lacoste J, Marcello M, Martins GG, Metcalf DJ, Mitchell CA, Moore J, Mueller T, Nelson MS, Ogg S, Onami S, Palmer AL, Paul-Gilloteaux P, Pimentel JA, Plantard L, Podder S, Rexhepaj E, Royon A, Saari MA, Schapman D, Schoonderwoert V, Schroth-Diez B, Schwartz S, Shaw M, Spitaler M, Stoeckl MT, Sudar D, Teillon J, Terjung S, Thuenauer R, Wilms CD, Wright GD, Nitschke R (2021) QUAREP-LiMi: A community-driven initiative to establish guidelines for quality assessment and reproducibility for instruments and images in light microscopy. J Microsc. 284(1):56-73. doi: 10.1111/jmi.13041.

Pfitzner S, Bosse JB, Hofmann-Sieber H, Flomm F, Reimer R, Dobner T, Grünewald K, Franken LE (2021) Human Adenovirus Type 5 Infection Leads to Nuclear Envelope Destabilization and Membrane Permeability Independently of Adenovirus Death Protein. Int J Mol Sci. 22(23):13034. doi: 10.3390/ijms222313034.

2020

Vollmer B, Pražák V, Vasishtan D, Jefferys EE, Hernandez-Duran A, Vallbracht M, Klupp BG, Mettenleiter TC, Backovic M, Rey FA, Topf M, Grünewald K (2020) The prefusion structure of herpes simplex virus glycoprotein B. Sci Adv. 6(39):eabc1726. doi: 10.1126/sciadv.abc1726.

Dietzel S, Ferrando-May E, Fried H, Kukat C, Naumann A, Nitschke R, Pasierbek P, Peychl J, Rasse TM, Schroth-Diez B, Stöckl MT, Terjung S, Thuenauer R, Tulok S, Weidtkamp-Peters S; German BioImaging-Society for Microscopy and Image Analysis (2020) A Joint Action in Times of Pandemic: The German BioImaging Recommendations for Operating Imaging Core Facilities During the SARS-Cov-2 Emergency. Cytometry A. 97(9):882-886. doi: 10.1002/cyto.a.24178.

Schubert T, Sych T, Madl J, Xu M, Omidvar R, Patalag LJ, Ries A, Kettelhoit K, Brandel A, Mely Y, Steinem C, Werz DB, Thuenauer R, Römer W (2020) Differential recognition of lipid domains by two Gb3-binding lectins. Sci Rep. 10(1):9752. doi: 10.1038/s41598-020-66522-8.

Thuenauer R, Landi A, Trefzer A, Altmann S, Wehrum S, Eierhoff T, Diedrich B, Dengjel J, Nyström A, Imberty A, Römer W (2020) The Pseudomonas aeruginosa Lectin LecB Causes Integrin Internalization and Inhibits Epithelial Wound Healing. mBio. 11(2):e03260-19. doi: 10.1128/mBio.03260-19.

Lunelli M, Kamprad A, Bürger J, Mielke T, Spahn CMT, Kolbe M (2020) Cryo-EM structure of the Shigella type III needle complex. PLoS Pathog. 16(2):e1008263. doi: 10.1371/journal.ppat.1008263.

Burda PC, Crosskey T, Lauk K, Zurborg A, Söhnchen C, Liffner B, Wilcke L, Pietsch E, Strauss J, Jeffries CM, Svergun DI, Wilson DW, Wilmanns M, Gilberger TW (2020) Structure-Based Identification and Functional Characterization of a Lipocalin in the Malaria Parasite Plasmodium falciparum. Cell Rep. 31(12):107817. doi: 10.1016/j.celrep.2020.107817.

Del Toro D, Carrasquero-Ordaz MA, Chu A, Ruff T, Shahin M, Jackson VA, Chavent M, Berbeira-Santana M, Seyit-Bremer G, Brignani S, Kaufmann R, Lowe E, Klein R, Seiradake E (2020) Structural Basis of Teneurin-Latrophilin Interaction in Repulsive Guidance of Migrating Neurons. Cell. 180(2):323-339.e19. doi: 10.1016/j.cell.2019.12.014.

Geiger M, Brown C, Wichers JS, Strauss J, Lill A, Thuenauer R, Liffner B, Wilcke L, Lemcke S, Heincke D, Pazicky S, Bachmann A, Löw C, Wilson DW, Filarsky M, Burda PC, Zhang K, Junop M, Gilberger TW (2020) Structural Insights Into PfARO and Characterization of its Interaction With PfAIP. J Mol Biol. 432(4):878-896. doi: 10.1016/j.jmb.2019.12.024.

2019

Kampf LL, Schneider R, Gerstner L, Thünauer R, Chen M, Helmstädter M, Amar A, Onuchic-Whitford AC, Loza Munarriz R, Berdeli A, Müller D, Schrezenmeier E, Budde K, Mane S, Laricchia KM, Rehm HL, MacArthur DG, Lifton RP, Walz G, Römer W, Bergmann C, Hildebrandt F, Hermle T (2019) TBC1D8B Mutations Implicate RAB11-Dependent Vesicular Trafficking in the Pathogenesis of Nephrotic Syndrome. J Am Soc Nephrol. 30(12):2338-2353. doi: 10.1681/ASN.2019040414.

Wang C, Lunelli M, Zschieschang E, Bosse JB, Thuenauer R, Kolbe M (2019) Role of flagellar hydrogen bonding in Salmonella motility and flagellar polymorphic transition. Mol Microbiol. 112(5):1519-1530. doi: 10.1111/mmi.14377.

Wichers JS, Scholz JAM, Strauss J, Witt S, Lill A, Ehnold LI, Neupert N, Liffner B, Lühken R, Petter M, Lorenzen S, Wilson DW, Löw C, Lavazec C, Bruchhaus I, Tannich E, Gilberger TW, Bachmann A (2019) Dissecting the Gene Expression, Localization, Membrane Topology, and Function of the Plasmodium falciparum STEVOR Protein Family. mBio. 10(4):e01500-19. doi: 10.1128/mBio.01500-19