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ZEISS Apotome 3
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ZEISS Apotome 3​ Optical Sectioning in Fluorescence Imaging for Your Widefield Microscope​

With structured illumination, removal of out-of-focus light becomes simple and efficient. ZEISS Apotome 3 calculates your optical section from a number of images acquired with different grid positions. Get high-contrast images, even from thicker specimens, while your system remains just as easy to operate as always.

  • Brilliant optical sections​
  • Based on peer-reviewed algorithms
  • More structural information 
  • Free choice of light source and dyes​
Transgenic zebrafish larva. Courtesy of H. Reuter, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

Brilliant Optical Sections​

Even From Thick Specimens

Apotome 3 significantly increases the axial resolution compared to conventional fluorescence microscopy: You obtain optical sections that allow 3D rendering, even from thick specimens. Three grids of different geometries give you the best resolution for each objective. You can focus on your experiment as the ideal grid is automatically selected, always resulting in high-contrast optical sections.

 

Caption: Transgenic zebrafish larva. Courtesy of H. Reuter, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

Cortical neurons (left: Widefield; right: Apotome 3). Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.
Cortical neurons (left: Widefield; right: Apotome 3). Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

Peer-reviewed Algorithms

Linear Approaches for True Optical Sections

Software-based solutions require either prior knowledge of the​ sample (AI based methods) or rely on complex algorithms that​ have not been peer-reviewed. Users must trust that these​ black-box solutions only produce structures that are real. ZEISS Apotome 3 uses linear approaches and well documented algorithms, allowing you to calculate true, reliable optical sections.

Caption: Cortical neurons (left: Widefield; right: Apotome 3). Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

Cortical neurons. Image 1 - Widefield, image 2 - Apotome 3, image 3 - Apotome 3 + DCV

Cortical neurons

Cortical neurons. Image 1 - Widefield, image 2 - Apotome 3, image 3 - Apotome 3 + DCV Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.
Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

More Structural Information

Compare Widefield, Optical Sectioning, and Deconvolution

Improve your images even more by deconvolution, using an algorithm for structured illumination. While retaining all raw data, the system allows you to switch between widefield, optical section and deconvolved images for maximum flexibility and best comparability. The robust, easy to use deconvolution algorithms improve both lateral and axial resolution. Improved contrast and noise suppression let you better recognize the structures of the examined object.

Caption: Cortical neurons. Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

Free Choice of Light Source and Dyes

Free Choice of Light Source and Dyes

It’s Your Decision, Not the Technology’s

Your experiments often evolve in complexity and requirements. That’s why you need adaptable equipment. Use Apotome 3 with metal halide lamps, economic white light LEDs, or the gentle, multi-color Colibri illumination system. Whether you work with DAPI, Alexa488, Rhodamin, Cy5, or with vital dyes such as GFP or mCherry – Apotome 3 adapts to your fluorophores and light source, creating the sharp and brilliant images you expect.

Your Flexible Choice of Components

Customize Your Microscope by Combining Apotome 3 with the Accessories Required for Your Research
  • Microscope​

    Microscope​

    • Axio Observer series (inverted research microscope)​​
    • Axio Imager 2 series (upright research microscope)​​
    • Axio Zoom.V16 (zoom microscope)​​
    • Simple upgrading of existing systems
  • Recommended objective classes​

    Recommended objective classes​

    • C-Apochromat​​
    • Plan-Apochromat​​
    • EC Plan-Neofluar
  • Illumination: Colibri 7

    Illumination

    • Colibri 5 and 7 (LED)​​
    • Xylis LED (white light LED)​​
    • HBO (mercury vapor lamp)​​
    • HXP 120 C (metal halide)
  • Cameras: Axiocam 712 mono

    Cameras

    • Monochrome, low-noise ZEISS Axiocam​ camera models​​
    • Selected 3rd-party cameras

Your Insight into the Technology Behind It​

Technology behind it: Optical sectioning using structured illumination allows you to efficiently minimize out-of-focus light to create crisp images and 3D renderings.
Technology behind it. Optical sectioning using structured illumination allows you to efficiently minimize out-of-focus light to create crisp images and 3D renderings.

Light from outside the focal plane needs to be suppressed to extract the in-focus image information. Optical sectioning using structured illumination allows you to efficiently minimize out-of-focus light to create crisp images and 3D renderings.

Quantitative Optical Sectioning

True Optical Sections Using Structured Illumination

Light from outside the focal plane needs to be suppressed to extract the in-focus image information. Optical sectioning using structured illumination allows you to efficiently minimize out-of-focus light to create crisp images and 3D renderings.

A: Widefield image. B – D: Raw images acquired with different grid positions. E: Resulting image; out-of-focus light is efficiently removed by the structured illumination.
A: Widefield image. B – D: Raw images acquired with different grid positions. E: Resulting image; out-of-focus light is efficiently removed by the structured illumination.

A: Widefield image. B – D: Raw images acquired with different grid positions. E: Resulting image; out-of-focus light is efficiently removed by the structured illumination.

A: Widefield image. B – D: Raw images acquired with different grid positions. E: Resulting image; out-of-focus light is efficiently removed by the structured illumination.

The Apotome 3 Operation Principle

Apotome 3 uses a grid to generate a pattern of intensity differences. If out-of-focus light is present at a certain region of the sample, the grid becomes invisible. After the fluorescence of a grid position is acquired, the grid moves to the next position. A true optical section with higher contrast and resolution is calculated.

C. elegans, whole mount, green: GFP, blue: DAPI. Objective: Plan-Apochromat 20 ×/0.8.
C. elegans, whole mount, green: GFP, blue: DAPI. Objective: Plan-Apochromat 20 ×/0.8. Courtesy of Prof. Schnabel, T.U. Braunschweig, Germany.
Courtesy of Prof. Schnabel, T.U. Braunschweig, Germany.

C. elegans, whole mount, green: GFP, blue: DAPI. Objective: Plan-Apochromat 20 ×/0.8.

C. elegans, whole mount, green: GFP, blue: DAPI. Objective: Plan-Apochromat 20 ×/0.8. Courtesy of Prof. Schnabel, T.U. Braunschweig, Germany.

Optimal Section Volumes for Your Sample​

No matter which magnification you are using – Apotome 3 automatically places the optimum grid in the beam path. ​

A: Emission light from out-of-focus areas is detected. Contrast and resolution are reduced. B: Reduction of unwanted background fluorescence increases with the grid frequency. The optical section becomes thinner. C: Image information from outside of the focal plane is suppressed. This improves contrast and resolution of the optical section. D: “Low grid” delivers the optimal section thickness in this example. Images of this type are particularly suitable for 3D analyses and the processing with rendering software.

3D rendering of cortical neurons stained for DNA and microtubules. Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

ZEISS Apotome 3 at Work​

Application Examples

     

See more application examples
ZEISS Apotome 3 at Work
Conventional Fluorescence - Drosophila neurons, blue: DAPI, yellow: GFP. Objective: Plan-Apochromat 20×/0.8. Courtesy of M. Koch, Molecular and Developmental Genetics, University of Leuven, Belgium.​
Apotome 3​ - Drosophila neurons, blue: DAPI, yellow: GFP. Objective: Plan-Apochromat 20×/0.8. Courtesy of M. Koch, Molecular and Developmental Genetics, University of Leuven, Belgium.​
Conventional Fluorescence | Apotome 3​

Drosophila Neurons​

Drosophila neurons, blue: DAPI, yellow: GFP. Objective: Plan-Apochromat 20×/0.8. Courtesy of M. Koch, Molecular and Developmental Genetics, University of Leuven, Belgium.​

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Drosophila embryo

Drosophila embryo, green: HRP, red: glia marker, 100 µm Z-stack. Courtesy of C. Klämbt, Institute for Neurobiology, University of Münster, Germany.

Comparison of a widefield image and 3D rendering of cortical neurons stained for DNA and microtubules. Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.
Comparison of a widefield image and 3D rendering of cortical neurons stained for DNA and microtubules. Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.
Widefield | Apotome 3

Cortical Neurons

Comparison of a widefield image and 3D rendering of cortical neurons stained for DNA and microtubules. Courtesy of L. Behrendt, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.

Left to right: Widefield, Apotome 3, Apotome 3 + Deconvolution​

Lotus Japonicus Root

Left to right: Widefield, Apotome 3, Apotome 3 + Deconvolution​

Left to right: Widefield, Apotome 3, Apotome 3 + Deconvolution​

Left to right: Widefield, Apotome 3, Apotome 3 + Deconvolution​

Lotus Japonicus Root

Autofluorescence of a Lotus Japonicus root infected with symbiotic bacteria stained with mcherry. Courtesy of F. A. Ditengou, University of Freiburg, Germany.

Top to bottom: Widefield, Apotome 3, Apotome 3 + Deconvolution

Transgenic Zebrafish Larvae​

Top to bottom: Widefield, Apotome 3, Apotome 3 + Deconvolution

Top to bottom: Widefield, Apotome 3, Apotome 3 + Deconvolution

Top to bottom: Widefield, Apotome 3, Apotome 3 + Deconvolution

Transgenic Zebrafish Larvae​

Transgenic zebrafish larvae at 4 days post fertilization staining for: Glial fibrillary acidic protein, acetylated Tubulin, GFP and DNA. Embedded in 1.2% low melt agarose. Courtesy of H. Reuter, Leibniz-Institute on Aging – Fritz-Lipmann-Institut e.V. (FLI), Germany.​

Typical Applications

Task
ZEISS Apotome 3 Function
Cell Culture
2D imaging
✓ 2D single images
Fast imaging of a 2D image
✓ Optical section available online on the monitor
Reliable detection of the marker even with strong background fluorescence
✓ Automatic grid selection for optimum contrast with each objective
Combination of multiple contrast techniques
✓ Any combination of fluorescence channels, brightfield, DIC and phase contrast
✓ Individual configuration of each fluorescence channel as an optical section or widefield image
Live Cell Imaging
Reduction of phototoxicity
✓ Particularly low phototoxicity in combination with LED illumination and high-sensitive cameras like ZEISS Axiocams
Time-lapse images
✓ Depending on the exposure time, up to three images per second
✓ Doubling of the frame rate with “burst mode”
Vibratome Sections, Histological Samples
3D imaging
✓ Automatic selection of the optimum grid for each objective
Modification of the optical section thickness
✓ Grid freely selectable depending on the specimen
Penetration depth
✓ Depending on the optical density of the tissue
3D reconstruction
✓ Rendering of the image stack via integrated software function
✓ Automatic transfer of the parameters of the individual fluorescence channels
Quantitative analysis
✓ Reproducible size measurements through automatic system calibration
Whole Mounts
3D imaging
✓ Multi Channel, Z Stack and Time Lapse, Deconvolution, images in raw data mode, 3D Rendering
Large area imaging
✓ Automatic acquisition of large sections using Tiles & Positions

Articles Related to Apotome 3

Imaging of Living Samples - New Technologies for Fast and Gentle Image Acquisition
11 June 2020

Imaging of Living Samples

Webinar - 35 min watch
Dr. Kalliopi Arkoudi
Sample Preparation for Fluorescence Microscope – Basic Introduction for Better Imaging Results
10 June 2020

Sample Preparation for Fluorescence Microscope

Webinar - 40 min watch
Yibing Zhao
Deconvolution: Basics and Practice
13 December 2021

Cell Census to Synapse: Capture the Big Picture and the Details with ZEISS Axio Observer 7

Webinar - 24 min watch
Colleen Manning

Related Applications

Cell Imaging with Fluorescence Microscopy

With different technologies available that balance sensitivity, resolution and fluorophore requirements, explore your options from widefield to deconvolution, confocal to super-resolution.

Looking Deep Into Chemically Cleared Brain & Thick Tissues

With multiple systems optimized for cleared tissues, find the right technology that balances sample size and the level of detail you wish to resolve.

Downloads

    • Preview image of ZEISS Apotome 3

      ZEISS Apotome 3

      Optical sectioning in fluorescence imaging for your widefield microscope
      File size: 3 MB
      Download
    • Preview image of ZEISS Apotome 3 - Flyer

      ZEISS Apotome 3 - Flyer

      Hardware-based, Quantitative Optical Sectioning with Well Documented Algorithms
      File size: 1 MB
      Download

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