Widefield - ZEISS Elyra 7 Lattice SIM² (Irchel) | Center for Microscopy and Image Analysis

ZEISS Elyra 7 Lattice SIM²

The Zeiss Elyra 7 with different super-resolution modalities, allows detailed visualization of samples beyond the diffraction limit of conventional light microscopy. With its 2 fast sCMOS cameras, this system is able to generate multiple color data simultaneously at incredibly high frame rates.

Responsible Person	responsible person
Location	University Zurich, Irchel Campus, Room Y42-H-79.
Training Request	Follow this link to apply for an introduction to the microscope

The Elyra 7 features the following modalities:

Widefield, DIC

Widefield (WF) mode (sample illumination with arc lamp), Laser WF mode (sample illumination with laser)

Lattice SIM

Structured Illumination Microscopy, allowing fast and gentle super-resolution imaging (~120 nm in xy and ~300 nm in z) in 3 dimensions.

Lateral resolution (XY): 120 nm, axial resolution (Z): 300 nm (typical experimental FWHM values with objective lens Plan-Apochromat 63× / 1.40 Oil DIC, subresolution beads of 40 nm diameter and excitation at 488 nm)

Lattice SIM²

The new SIM² module doubles the conventional SIM resolution and achieves up to ~60 nmlaterally and ~200 nm axially. The Lattice SIM illumination allows for a better signal to noise ratio, more gentle imaging and can reach up to 255 fps during time lapse acquisition, well suited to capture highly dynamic biological processes in live samples. It further allows deeper imaging of samples up to around 70 µm thickness in comparison to only 20 µm of sample thickness in classical SIM. The speed of this imaging modality is well suited to capturing dynamic biological processes in live samples (significantly faster than regular laser scanning confocal).

Lateral resolution (XY): down to 60 nm, axial resolution (Z): down to 200 nm (typical experimental FWHM values with objective lens Plan-Apochromat 63× / 1.40 Oil DIC, subresolution beads of 40 nm diameter and excitation at 488 nm; Resolution is sample and SNR dependent.)

Apotome

Grid-based optical sectioning to create highly contrasted images with high lateral and axial resolution. The Apotome mode is also amenable to SIM2 processing, allowing for high speed acquisition with high contrast, high resolution and low phototoxicity. In combination with SIM² module, apotome imaging can get lateral rsoltuion down to ~110 nm and axial resolution to ~300 nm. ApoTome imaging is also faster than classic SIM and laser scanning microscopes, allowing for time lapse imaging of faster cellular/tissue dynamics.

Lateral resolution (XY) of 140 nm, axial resolution (Z) of 275 nm for 40×; lateral resolution (XY) of 285 nm, axial resolution (Z) of 550 nm for 20×; lateral resolution (XY) of 710 nm, axial resolution (Z) of 1300 nm for 10×

SMLM

Single-molecule localization microscopy like dSTORM, PALM and PAINT allowing resolution down to ~20-30 nm in xy and ~50-80 nm in z.

HILO & TIRF

Highly inclined and laminated optical sheet (HILO) and total interbal reflection illumination (TIRF).

Technical Specifications

Microscope body

inverted widefield microscope (Axio Observer 7 SR RP Stativ for ELYRA 7)
Piezo Stage for fast, smooth, repeatable motion over distances of up to 500 microns with sub-nanometer resolution
"online" super-resolution image projection - see the emerging image during aquisition
Large field of view - 1280 x 1280 pixels, 82um FOV (at 63x)
Definitiv Focus
Incubator XL dark and top stage incubation (full environmental chamber, temperature and CO2 control)

Light Sources and Lasers

Halogen lamp for transmitted light
External fluorescence lamp HXP 120 V
Solid state diode lasers:
- 405 nm (50 mW)
- 488 nm (500mW)
- 561 nm (500 mW)
- 642 nm (500 mW)

Camera System

2x pco.edge 4.2 sCMOS (duo link for dual camera)
16 bit cooled monochrome camera
2048 x 2048 pixel (6.5 x 6.5 um pixel size) back-illuminated sCMOS
QE: 82 %
dynamic range 15 bit

Environmental control

TempModul S1 - reliable control of of temperature up to 0.1°C accuracy
CO2-Modul S1 - allows for a stable concentration between 1-8% (in 0.1% steps)

Accessories

KM Frame for well plates
Click-In for petri dishes (35 mm and 60 mm)
Click-In for standard object slides, µ-Slides by Ibidi, Nunc Lab-Tek Slides and Chambers by Thermo Scientific

Available Optics

Name	Magnification	NA	Immersion	WD (mm)	Application
ZEISS Plan-Neofluar	2.5x	0.085	Air	8.8	Widefield
ZEISS Plan-Neofluar	10x	0.3	Air	5.2	Widefield, Apotome mode
ZEISS Plan-Apochromat	20x	0.8	Air	0.55	Widefield, Apotome mode
ZEISS Plan-Apochromat	40x	1.4	Oil	0.13	Widefield, DIC, Apotome mode
ZEISS Plan-Apochromat	63x	1.4	Oil	0.19	Widefield, DIC, SIM
ZEISS Plan-Apochromat TIRF	63x	1.46	Oil	0.1	Widefield, SMLM, TIRF, HILO

Available Filters

Mode	Main beam splitter	Emission Filter
FSet 77 HE (3 colour triple filter green, red, far red)	-	-
Transmitted Light	-	-
TIRF/Apotome/SIM/SMLM	405/488/561/642	LBF 405/488/561/642
TIRF/Apotome/SIM/SMLM	405/488/561/642	BP 420-480 / BP 495-525 / LP 655
TIRF/Apotome/SIM/SMLM	405/488/561/642	BP 495-550 / BP 570-620
TIRF/Apotome/SIM/SMLM	405/488/561/642	BP 420-480 / LP 655

: Zoom

Sample requirements

SMLM / TIRF

SMLM and TIRF often have special requirements in terms of fluorescence dyes and buffer systems. Be sure to contact the responsible persons before planning your experiments.

SIM

Coverslips must be #1.5H size thickness and made with high precision glass (170 ±5μm), features required to achieve “super resolution” precision.
All mounting media must have a RI of >1.4. We recommend ProLong Diamond / SlowFade Glass (Invitrogen/Life Tech).
All common fluorescent proteins and dyes are suitable for SIM, provided that they are compatible with the following available laser lines: 405, 488, 561, and 642nm.
Since the final quality of the reconstructed SIM image is dependent on the signal-to-noise ratio, samples that have been highly optimized previously to have low background with strong fluorescent labeling will yield the best images.

Acquisition speed (Lattice SIM)

17 SIM image frames per second at 512 × 512 px resolution and 1 ms exposure time (15 phase images per one SIM image)
19 SIM image frames per second at 512 × 512 px resolution and 1 ms exposure time (13 phase images per one SIM image)
28 SIM image frames per second at 512 × 512 px resolution and 1 ms exposure time (9 phase images per one SIM image)

Acquisition speed (SIM Apotome mode)

51 sectioned frames per second at 512 × 512 px resolution and 1 ms exposure time (camera limited) (5 phase images per one sectioned image);
85 sectioned frames per second at 512 × 512 px resolution and 1 ms exposure time (camera limited) (3 phase images per one sectioned image);

Leap mode and Burst mode

Burst mode: Processing for 2D time series data sets for Lattice SIM and Aptotome mode to increase effective frame rates by a factor of 15 and 5, respectively
- Max. 255 image frames per second at 512 × 512 px resolution and 1 ms exposure time are available for 2D data after Burst processing.Leap and Burst modes are combinable with both the Lattice SIM and SIM Apotome.
Leap mode increases the frame rate by a factor of 3 for 3D image acquisition.