We develop novel optical methods and instrumentation for the observation of biological phenomena with the highest fidelity. In particular, we focus on super resolution fluorescence imaging and single molecule tracking implementations that offer outstanding spatial and/or temporal resolution. By bringing together expertise in optics, electronics, data processing, labelling and photophysics, we develop solutions to transversal obstacles in biology and biophysics such as quantification of protein clusters and complexes, and identification of protein interaction and dynamics.


We are hosted and funded by the Research Institute of Molecular Pathology within the Vienna BioCenter and also recieve funding from the European Research Council.  

Super resolution microscopy


Fluorescence microscopy is an invaluable tool for exploring the structure and function of biological processes. It provides high specificity and contrast for the observation of cellular components (DNA, RNA, proteins, lipids, etc.) tagged with fluorescent molecules in a minimally invasive fashion, even allowing the study of live specimens. The spatial resolution of classical fluorescence microscopy is limited tohundreds of nanometers due to the diffraction of light; however, higher resolutions were unlocked with the development of the so-called super-resolution methodologies (stimulated emission depletion (STED) microscopy, photo-activated localization microscopy/stochastic optical reconstruction microscopy (PALM/STORM), among others)


In the last decade, achieving resolutions in the order of 10 nm to 100 nm became routine and has revealed details of subcellular organelles and new structures as well as ultrastructural anatomy in tissue, granting the Nobel Prize in Chemistry in 2014 to the developers of super resolution. Despite this revolution, the development of an ultimate microscope –revealing the precise nanometric location of all molecules of interest at all times without affecting the sample under study– remains elusive. Among several adversities, the photon budget of fluorescent probes is a fundamental bound for the trade-off between spatial and time resolution.



Advanced light microscopy and biophysics


How can light-based methods provide maximal information on biological processes? What are the limits to the information we can extract? How deep and fast can we look? How can we use extreme resolution to answer concrete questions? What answers can multimodal approaches deliver?


The development of novel optical methods like MINFLUX (see below) shows that the answers to these questions remain unexplored or incomplete, and that there is much to gain from synergistically combining expertise from diverse areas like optics, electronics, statistics, chemistry and biology. By solving transverse methodological challenges, we strive to push forward the maturation of light microscopy and profoundly influence life sciences along the way. 



To overcome this, maximally informative luminescence excitation or MINFLUX, merges elements of information theory with (i) the single-emitter nature of PALM/STORM (fig. 1A) and (ii) the beam geometries typically used in STED (e.g the so-called “doughnut” beam). This technique (fig. 1 B–E) has shown that the information that each photon contains on the location of its emitter is a flexible quantity and that it can be dramatically increased in imaging and tracking applications (fig. 2B–D). Therefore, a given localization precision can be obtained by using much fewer photons (e.g. 20 times) than in conventional centroid-localization techniques, such as PALM and STORM.

gallery/imp webstie - figure 1 v1

Figure 1. (A) Principle of PALM/STORM, where the resolution depends on the wavelength, numerical aperture and number of collected photons. (B) MINFLUX depicted as a ruler, where emitters are located from a sequence of exposures to tailored light patterns. The resolution now depends on the size of the ruler (L, separation between sequential excitations), instead of the wavelength. (C) A MINFLUX scheme in 2D with multiple excitations with doughnut-shaped beams, each coloured dot is the center of an exposure. (D) A MINFLUX scheme in 3D, with a beam created by top-hat wavefront shaping. (E) Iterative MINFLUX, where successive approximations to the location of the molecule produce a localization that surpasses the typical N-1/2 dependence..

gallery/imp webstie - figure 2 v2

Figure 2. (A) MINFLUX nanoscopy by sequentially localizing individually blinking fluorophores; a DNA-origami arrangement is imaged. (B) MINFLUX tracking of a DNA-origami flipping device, a fluorophore is followed with 2.5 nm precision every ~0.5 ms. (C) MINFLUX tracking of the small ribosomal subunit protein in living E. coli. (D) Iterative MINFLUX imaging of nuclear pore complex in 3D (fixed) and 2D (live, with comparison to classical SMLM), and of the PSD-95 protein at a neuronal synapse.


Group Leader

gallery/eva-maria wiedemann

Eva-Maria Wiedemann

Research Assistant

  • Since 2020 Research Group Leader at IMP, Vienna, Austria
  • 2020–2024 Starting Grant from the European Research Council.
  • 2012–2019 Post Doctoral Researcher at MPI for Biophysical Chemistry, Göttingen, Germany
  • 2007–2012 PhD at University of Buenos Aires, Argentina
  • 2002–2007 Electrical Engineering at University of Buenos Aires, Argentina

Quick CV

gallery/Mehrta Shirzadian



PhD Candidate

gallery/Max Geismann 2



PhD Candidate

Open Positions







Call details



Positions calls are now open (last update 10.02.2020)


Visit the Vienna BioCenter PhD program for more information on PhD positions. 

PhD Application Deadlines:

Winter Selection: September 15th - November 15th.

Summer Selection: March 1st - April 15th.

Meet us!






27 Aug 2020

Francsico Balzarotti

Invited Talk

MINFLUX: Super Resolution at the molecular scale

26-28 Aug


Francisco Balzarotti

Invited Talk



Single Molecule Localization Symposium

Lausanne, Switzerland

23-28 Aug


Francisco Balzarotti

Invited Talk

Super Resolution Microscopy at the Molecular Scale


European Microscopy Conference 2020

Copenhagen, Denmark

11-16 Jul






05 Mar





MINFLUX: Next generation Super Resolution Microscopy and Single Molecule Tracking

Seminar Series: Modern Concepts in Structural Biology
VBC5 - Vienna BioCenter (Location)

02 Feb


Francisco Balzarotti

Invited Talk

MINFLUX: Achieving the ultimate resolution limit in fluorescence microscopy 

Photonics West BIOS

San Francisco, USA

06 Jan


Francisco Balzarotti


Isotropic 3D molecular resolution in cells with MINFLUX nanoscopy 

Find us at the following events: 

  • NEW! Multicolor 3D MINFLUX nanoscopy of mitochondrial MICOS proteins
    Pape, J.K., Stephan, T., Balzarotti, F., Büchner, R., Lange, F., Riedel, D., Jakobs, S., Hell, S.W.
    PNAS (2020)

  • NEW! MICOS assembly controls mitochondrial inner membrane remodeling and crista junction redistribution to mediate cristae formation
    Stephan, T., Brüser, C., Deckers, M., Steyer, A.M., Balzarotti, F., Barbot, M., Behr, T.S., Heim, G., Hübner, W., Ilgen, P., Lange, F., Pacheu-Grau, D., Pape, J.K., Stoldt, S., Huser, T., Hell, S.W., Möbius, W., Rehling, P., Riedel, D., Jakobs, S.
    The EMBO Journal n/a, e104105. (2020)

  • NEW! MINFLUX nanoscopy delivers 3D multicolor nanometer resolution in cells
    Gwosch, K.C., Pape, J.K., Balzarotti, F., Hoess, P., Ellenberg, J., Ries, J., Hell, S.W.
    Nat. Methods 17, 217-224 (2020) | bioRxiv 734251
  • MINFLUX monitors rapid molecular jumps with superior spatiotemporal resolution
    Eilers, Y., Ta, H., Gwosch, K.C., Balzarotti, F., Hell, S.W. 
    PNAS USA 201801672 (2018)
  • Nanometer resolution imaging and tracking of fluorescent molecules with minimal photon fluxes
    Balzarotti, F., Eilers, Y., Gwosch, K.C., Gynnå, A.H., Westphal, V., Stefani, F.D., Elf, J., Hell, S.W. 
    Science 355, 606–612 (2017) | arXiv:1611.03401
  • Ultrastructural anatomy of nodes of Ranvier in the peripheral nervous system as revealed by STED microscopy
    D’Este, E., Kamin, D., Balzarotti, F., Hell, S.W.
    PNAS USA 201619553 (2016)
  • STED nanoscopy with wavelengths at the emission maximum
    Bordenave, M.D., Balzarotti, F., Stefani, F.D., Hell, S.W.
    Journal of Physics D: Applied Physics 49, 365102 (2016)
  • Dual Channel RESOLFT Nanoscopy by Using Fluorescent State Kinetics
    Testa, I., D’Este, E., Urban, N.T., Balzarotti, F., Hell, S.W.
    Nano Letters 15, 103–106 (2015)
  • Plasmonics Meets Far-Field Optical Nanoscopy
    Balzarotti, F., Stefani, F.D.
    ACS Nano 6, 4580–4584 (2012)


Balzarotti Lab 

Research Institute for Molecular Pathology

Campus-Vienna-BioCenter 1

1030, Vienna, Austria