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STED for malaria research: STED nanoscopy of merozoites invading erythrocytes. 3D STED image showing overlays of RON4 (magenta) with the proteins PfRh5 (left, green), PfRipr (middle, green), PfCyRPA (right, green). Nuclear staining with DAPI is shown in blue, and erythrocytic membrane is shown in gray. Scale bar: 1 µm. Image courtesy: Jennifer Volz, Alan Cowman, Walter and Eliza Hall Institute of Medical Research, Australia, and Marko Lampe, EMBL, Germany.

Nanoscopy continues revolutionizing the study of subcellular architecture and dynamics

The Nobel Prize in Chemistry 2014 was awarded “for the development of super-resolved fluorescence microscopy”. This groundbreaking work brought optical microscopy to a new era of fluorescence imaging beyond the diffraction limit, enabling scientists to observe molecules at the nano-level. Today, nanoscopy continues revolutionizing the study of subcellular architecture and dynamics.

STED on STELLARIS: Super-resolution for the life sciences

Combined with the STELLARIS platform, STED is a powerful and highly-versatile modality delivering cutting-edge multicolor, deep and live-cell nanoscopy results for your research.

  • τ-STED advances nanoscopy by enabling demanding super-resolution applications with gentle live cell imaging, fast acquisition, and excellent image quality.
  • Flexible nanoscopy at its fastest, thanks to the combination of fast white light laser (WLL) excitation, high speed resonant scanning and the excellent dynamic range of the STELLARIS new Power HyD detector family.
  • Up to 3 STED lines (592, 660, and 775 nm) combined with the high-end STED-specialized objective lenses open up the full spectrum of visible light for deep and live cell applications.
  • Fast auto-beam alignment for stability and reliability, protecting your sample from unnecessary light irradiation.
  • From confocal to STED fast and within seconds, thanks to the intelligence of the brand-new ImageCompass user interface.
STED for Immunology: Living T cell in suspension. 3D reconstruction of confocal and STED stacks. Image courtesy: Marco Fritsche, Mathias Clausen and Christian Eggeling, MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, UK.

τ-STED: Nanoscopy meets Lifetime

τ-STED, exclusively from Leica Microsystems, advances nanoscopy for demanding applications. This is possible thanks to the unique combination of STED, the ultra-fast Power HyD detector family and FLIM as provided by FALCON.

τ-STED identifies the signal from the STED process and differentiates it from background using fluorescence lifetime in a novel way. It decodes the fluorescence lifetime gradient to deliver the desired resolution at dramatically lower light dose, and to push the resolution beyond conventional STED. The results are high quality images at resolutions better than 30 nm (lateral) and 100 nm (axial).* 

τ-STED works for 2D and 3D multicolor STED, in live and fixed specimens. The lower excitation and STED light dose protect your sample, extend time-lapse experiments and enable large volume imaging. And with FALCON, the raw data is always available for validation and quantification.

*depending on sample and fluorophore. 

STED for Developmental Biology: Single molecule in-situ hybridization (smFISH) of RNA in Drosophila embryo whole-mount preparation. Probes are directly labeled (no signal amplification). Top: Three color t-STED 775 captures the signal from hb CalFluor 610 (cyan), gt Quasar 670 (green), eve Quasar 705 (magenta). Bottom: confocal overview of the Drosophila embryo preparation. Sample courtesy: Tom Pettini, University of Manchester, UK. 

Push the resolution while protecting your sample

τ-STED uses the lifetime information contained in every STED experiment to eliminate background  and map the STED response of the fluorophore in real time. Access to this information allows you to increase image quality (expressed as signal/noise ratio) and resolution without having to resort to harsh imaging conditions.

τ-STED benefits from the flexibility of the STED platform: WLL excitation and STED effect all over the spectrum, optimized for your fluorophore of choice.

τ-STED 775 imaging of GATTA-Bead R 23 nm. Scale bar: 1um.

Get the super-resolution and quality you need at low light dose

When you look for low illumination strategies for your STED experiment, there is a key question: How low is low? The answer is far from trivial and from universally fixed values because, if you think about this, it depends on your sample, your label, and application.

We will not promise arbitrary light dose reductions that do not relate to your experiment. Instead, we can show you that τ-STED significantly reduces the light dose from the STED and the excitation in your sample. τ-STED eliminates the background and reaches resolutions that otherwise require significantly higher compensation dosage and loss of quantification capabilities.

τ-STED 660 on COS7 cells immuno-stained for nuclear pores (NUPs). The primary antibody recognizes several proteins from the nuclear pore basket, thus the point-like structures observed. Only 2% of STED light reveals a great amount of additional detail.
Confocal
τ-STED

STED for kidney disease research: ​​Two color, 2D STED (top) and 3D STED (bottom) imaging of non-cleared mouse kidney section immunostained for synaptopodin (green) and nephrin (magenta). Sample courtesy: Victor Puelles, Milagros Wong and Jan Czogalla, Universitätsklinikum Hamburg-Eppendorf, Hamburg.

τ-STED provides photoprotection in all dimensions

τ-STED works for multicolor applications. You can use the best dyes in the red region of the spectrum, go for fluorescent proteins in the green, or exploit novel fluorogenic probes in the orange. STED on STELLARIS is equipped with our WLL with AOBS to excite your sample where needed, and enables you to choose STED 775, 592, or our exclusive 660 laser lines.

Gentle nanoscopy for extended time-lapse experiments

The lower excitation and STED light dose translates into protection for the specimen, empowering longer time-lapse experiments (more frames), or larger volume imaging, without sacrificing spatial resolution.

Gentle live cell τ-STED 592 on mammalian cells stably expressing LifeAct-mNeonGreen. Scale bar: 5mm. The timelapse here shows 230 frames @1 fps. Sample courtesy: Max Heydasch, University of Bern, Switzerland.

Multicolor STED nanoscopy

Cover the whole spectrum of visible light

Multicolor applications give access to detailed information about the relationships of various structures. Colocalization studies are routinely performed with STED. The multiple STED laser lines of STED at 592, 660, and 775 nm, cover the whole spectrum of visible light and give access to many applicable fluorophores.

More colors make the difference!

The White Light Laser, the AOBS and the tunable Power HyD family of detectors synergistically enable imaging any kind of fluorophore combination and give you the highest flexibility for your multicolor experiment.

STED means for me: Seeing the essential details!

Prof. Stephan Sigrist, FU Berlin, Germany

Three color STED imaging of Cos 7 cells stained with antibodies against NUP153 (green), TOM20 (magenta), and with SiR–actin (gray). Confocal (top) and STED (bottom). Scale bar: 2μm.
Sample courtesy: Urs Ziegler and Jana Doehner, ZMB, University of Zurich, Switzerland.

Nanoscopy empowered by cutting-edge optics​

When it comes to nanoscopy resolution, it is crucial to start with the best optics. The chromatic correction of our STED WHITE Class of objective lenses ensures optimal overlay of excitation and STED PSF in z all over the visible spectral range.

We have developed three STED specialized objective lenses for daily, deep, and live cell nanoscopy.

Among our STED WHITE lenses, the STED WHITE 100x oil lens delivers the highest numerical aperture (NA) and resolution, and performs excellently for standard fixed samples. For deep STED nanoscopy experiments, the STED WHITE 93x Glycerol objective lens is the lens of choice. The motorized correction collar adapts precisely and swiftly to cover glass thickness, temperature changes and specimen inhomogeneity, in 2D and 3D. 

The STED WHITE 86x Water lens is dedicated to water-based experiments. The motCORR technology makes this lens a powerful companion in live cell and STED-FCS applications.​

The 93x Glycerol lens is a must, everybody should have it!

Yasushi Okada, RIKEN Quantitative Biology Center, Japan 

STED WHITE Class dedicated lenses for STED imaging. Top: Chromatic aberration of STED WHITE lenses. Depth of field depicted in gray dashed line. Bottom: STED WHITE oil, and glycerol and water lenses equipped with motCORR technology.

STED in the 3rd dimension

You can choose between best lateral resolution, best axial resolution or anything in between to get optimal results. Ultra-thin optical sections reveal unknown details. STED combined with the power of STELLARIS enable you to match the resolution of your microscope in all dimensions to your question and specimen.

Adding the third dimension and an additional STED line to STED imaging allows us to see things that were impossible to see before.

Dr. Timo Zimmermann, Center for Genomic Regulation, Barcelona, Spain

STED for kidney disease research: 3D STED 775 deep nanoscopy of glomerulus in cleared kidney tissue immunostained for nephrin. Scale bar: 10 μm, z-depth color coded. Sample courtesy: David Unnersjö Jess, KTH, Stockholm, Sweden.

Select your resolution

Resolution improvements are accomplished by allocating the light to the two STED pathways: the best lateral resolution is achieved by the classic vortex donut, the best axial resolution by the z donut. It can also be optimized for the smallest focal volume.

Investigate molecular dynamics with STED FCS

STED provides a powerful tool to continuously tune the size of the observation area down to the nanoscale. For this reason, STED-FCS (fluorescence correlation spectroscopy) reports on the diffusion time, concentration, and binding constant of molecules even at higher concentrations (> 100 nM) than those possible with confocal FCS approaches.

STED-FCS can be combined with fluorescence lifetime imaging (FLIM) and related cross-correlation techniques (STED-FLCS, STED-FLCCS) enabling the study of different molecules whose fluorescence spectra may overlap.

STED calibration performed on lipid bilayers prepared with KK114. The autocorrelation curves obtained at increasing STED intensities (0%, 10%, 20%, 50% and 100%) are normalized at 1 μs. The shift of the autocorrelation curves indicates a decrease in the effective observation volume with STED. Courtesy of Christian Eggeling, University of Oxford, UK.

Software solutions for STED – know your resolution

STED on STELLARIS is fully integrated into the intuitive LAS X software platform from Leica, ensuring ease of use and fast user training. Smart guidance powered by our brand-new ImageCompass user interface optimizes settings for fast confocal and τ-STED acquisition and visualization in 2D & 3D.

Our intelligent STED automatic alignment is accomplished with a single click and protects your sample from laser light exposure. The powerful LAS X Navigator software produces rapid large-area overview images to provide context and rapid identification of regions for more detailed subsequent STED examination.

New ImageCompass user interface for STED on STELLARIS.

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