Microscopy | Universitätsklinikum Freiburg

Microscopy Trainings

Content of the Microscopy Trainings

Please note: There are several different microscopy systems available which need selective introductions before usage.

You will learn:

Startup operations, operation and shutdown procedures
Sample preparation
Sample acquisition
Sample handling and disposal of GMOs according to the “Gentechnikgesetz (GenTG)”
Spectral overlap compensation
Operating corresponding software
Troubleshooting

If you would like to use our widefield, confocal or high throughput systems and have not yet received instruction on this instruments from us, please request a training by send us an email to facslab(ät)uniklinik-freiburg.de with your preferred date for the instruction. You will then receive an email from us with all details and information.

Confocal microscopy basic course (on LSM710/880):

Confocal Microscopy Induction: Course Syllabus

Systems: Zeiss LSM Series (880/900/980) with Airyscan Technology

I. Theoretical Foundation: The Confocal Principle

Physics of Confocality: Understanding the pinhole, optical sectioning, and the rejection of out-of-focus light.
The Light Path:
- Excitation: Laser lines and Acousto-Optical Tunable Filters (AOTF).
- Emission: Secondary Dichroic Mirrors (SDM) and spectral detection.
Resolution and Contrast:
- The relationship between Pinhole size (Airy Units) and Z-section thickness.
- Nyquist sampling criteria for XY and Z.
Airyscan Technology: Super-resolution concepts (1.7x improvement) and the 32-channel GaAsP detector array.

II. Hardware Overview & Start-up Procedures

The Axio Imager Stand: Introduction to the motorized widefield components (eyepieces, condenser, and internal light paths).
Objective Care: Proper use of immersion media (Oil, Water, Glycerol) and the critical "cleaning after use" protocol.
Safe Start-up Sequence:
1. Main power/Component switches.
2. Systems/PC initialization.
3. Laser activation (warm-up requirements).
Incubation (if applicable): Setting up the heating and CO₂modules for live-cell imaging.

III. Software Operation (ZEISS ZEN Blue/Black)

The Workflow Concept: Locate (Widefield) vs. Acquisition (Confocal) modes.
Image Setup:
- Configuring Smart Setup vs. Manual configuration.
- Detectors: Setting Gain (Master), Offset, and Laser Power to minimize phototoxicity and bleaching.
Advanced Acquisition Modules:
- Z-Stacks: Defining bounds and optimal step size.
- Tile Scanning: Stitching and focus surface mapping (Definite Focus).
- Time Lapse: Managing temporal resolution and focus drift.
Airyscan Processing: Introduction to the "Airyscan Processing" tab and 2D/3D SR reconstruction.

IV. Data Management & Best Practices

The .CZI Format: Why you should always save in the native Zeiss format to preserve metadata.
Storage Policy:
- Local vs. Server storage (D: Drive policy).
- Automatic deletion/cleanup cycles of the facility.
Metadata: How to retrieve acquisition settings from existing files for reproducibility.

V. System Shutdown & Quality Control

The "Next User" Rule: Checking the facility calendar before complete shutdown.
Cool-down Protocol: Proper laser venting and shutdown sequence to protect sensitive electronics.
Hardware Maintenance:
- Cleaning objectives with lens tissue and Sparkle/Ethanol.
- Leave the system on lower magnification objective.
Reporting: How to log hours and report technical issues or hardware errors.

At an individual on-site appointment for a maximum of 2 people each (duration in total maximum 90 min.), the formalities are then completed and, if possible, permission to use the equipment is granted.

Always watch all videos on this list to be prepared for the course!

The next dates for the on-site appointments are planned from 9:00 -12:00 on:

16.02.2026
02.03.2026
23.03.2026
20.04.2026
11.05.2026
08.06.2026
22.06.2026
06.07.2026
20.07.2026
03.08.2026
17.08.2026
31.08.2026
14.09.2026
28.09.2026
12.10.2026
26.10.2026
09.11.2026
23.11.2026
07.12.2026
21.12.2026
04.01.2027
18.01.2027
01.02.2027
15.02.2027
01.03.2027
to be continued

Other confocal systems advanced introduction

Zeiss LSM980

Confocal Microscopy Induction: Course Syllabus

Systems: Zeiss LSM Series (880/900/980) with Airyscan Technology

I. Theoretical Foundation: The Confocal Principle

Physics of Confocality: Understanding the pinhole, optical sectioning, and the rejection of out-of-focus light.
The Light Path:
- Excitation: Laser lines and Acousto-Optical Tunable Filters (AOTF).
- Emission: Secondary Dichroic Mirrors (SDM) and spectral detection.
Resolution and Contrast:
- The relationship between Pinhole size (Airy Units) and Z-section thickness.
- Nyquist sampling criteria for XY and Z.
Airyscan Technology: Super-resolution concepts (1.7x improvement) and the 32-channel GaAsP detector array.

II. Hardware Overview & Start-up Procedures

The Axio Imager Stand: Introduction to the motorized widefield components (eyepieces, condenser, and internal light paths).
Objective Care: Proper use of immersion media (Oil, Water, Glycerol) and the critical "cleaning after use" protocol.
Safe Start-up Sequence:
1. Main power/Component switches.
2. Systems/PC initialization.
3. Laser activation (warm-up requirements).
Incubation (if applicable): Setting up the heating and CO₂modules for live-cell imaging.

III. Software Operation (ZEISS ZEN Blue/Black)

The Workflow Concept: Locate (Widefield) vs. Acquisition (Confocal) modes.
Image Setup:
- Configuring Smart Setup vs. Manual configuration.
- Detectors: Setting Gain (Master), Offset, and Laser Power to minimize phototoxicity and bleaching.
Advanced Acquisition Modules:
- Z-Stacks: Defining bounds and optimal step size.
- Tile Scanning: Stitching and focus surface mapping (Definite Focus).
- Time Lapse: Managing temporal resolution and focus drift.
Airyscan Processing: Introduction to the "Airyscan Processing" tab and 2D/3D SR reconstruction.

IV. Data Management & Best Practices

The .CZI Format: Why you should always save in the native Zeiss format to preserve metadata.
Storage Policy:
- Local vs. Server storage (D: Drive policy).
- Automatic deletion/cleanup cycles of the facility.
Metadata: How to retrieve acquisition settings from existing files for reproducibility.

V. System Shutdown & Quality Control

The "Next User" Rule: Checking the facility calendar before complete shutdown.
Cool-down Protocol: Proper laser venting and shutdown sequence to protect sensitive electronics.
Hardware Maintenance:
- Cleaning objectives with lens tissue and Sparkle/Ethanol.
- Leave the system on lower magnification objective.
Reporting: How to log hours and report technical issues or hardware errors.

The next dates for the on-site appointments are planned from 9:00 -12:00 on:

09.02.2026
23.02.2026
09.03.2026
30.03.2026
13.04.2026
04.05.2026
18.05.2026
01.06.2026
15.06.2026
29.06.2026
13.07.2026
27.07.2026
10.08.2026
24.08.2026
07.09.2026
21.09.2026
05.10.2026
19.10.2026
02.11.2026
16.11.2026
30.11.2026
14.12.2026
28.12.2026
11.01.2027
25.01.2027
08.02.2027
22.02.2027
08.03.2027
to be continued

Zeiss CD7/LSM900

Confocal Microscopy Induction: Course Syllabus

Systems: Zeiss LSM Series (880/900/980) with Airyscan Technology

I. Theoretical Foundation: The Confocal Principle

Physics of Confocality: Understanding the pinhole, optical sectioning, and the rejection of out-of-focus light.
The Light Path:
- Excitation: Laser lines and Acousto-Optical Tunable Filters (AOTF).
- Emission: Secondary Dichroic Mirrors (SDM) and spectral detection.
Resolution and Contrast:
- The relationship between Pinhole size (Airy Units) and Z-section thickness.
- Nyquist sampling criteria for XY and Z.
Airyscan Technology: Super-resolution concepts (1.7x improvement) and the 32-channel GaAsP detector array.

II. Hardware Overview & Start-up Procedures

The Axio Imager Stand: Introduction to the motorized widefield components (eyepieces, condenser, and internal light paths).
Objective Care: Proper use of immersion media (Oil, Water, Glycerol) and the critical "cleaning after use" protocol.
Safe Start-up Sequence:
1. Main power/Component switches.
2. Systems/PC initialization.
3. Laser activation (warm-up requirements).
Incubation (if applicable): Setting up the heating and CO₂modules for live-cell imaging.

III. Software Operation (ZEISS ZEN Blue/Black)

The Workflow Concept: Locate (Widefield) vs. Acquisition (Confocal) modes.
Image Setup:
- Configuring Smart Setup vs. Manual configuration.
- Detectors: Setting Gain (Master), Offset, and Laser Power to minimize phototoxicity and bleaching.
Advanced Acquisition Modules:
- Z-Stacks: Defining bounds and optimal step size.
- Tile Scanning: Stitching and focus surface mapping (Definite Focus).
- Time Lapse: Managing temporal resolution and focus drift.
Airyscan Processing: Introduction to the "Airyscan Processing" tab and 2D/3D SR reconstruction.

IV. Data Management & Best Practices

The .CZI Format: Why you should always save in the native Zeiss format to preserve metadata.
Storage Policy:
- Local vs. Server storage (D: Drive policy).
- Automatic deletion/cleanup cycles of the facility.
Metadata: How to retrieve acquisition settings from existing files for reproducibility.

V. System Shutdown & Quality Control

The "Next User" Rule: Checking the facility calendar before complete shutdown.
Cool-down Protocol: Proper laser venting and shutdown sequence to protect sensitive electronics.
Hardware Maintenance:
- Cleaning objectives with lens tissue and Sparkle/Ethanol.
- Leave the system on lower magnification objective.
Reporting: How to log hours and report technical issues or hardware errors.

The next dates for the on-site appointments are planned from 9:00 -12:00 on:

11.02.2026
18.02.2026
11.03.2026
01.04.2026
15.04.2026
06.05.2026
20.05.2026
03.06.2026
17.06.2026
01.07.2026
15.07.2026
29.07.2026
12.08.2026
26.08.2026
09.09.2026
23.09.2026
07.10.2026
21.10.2026
04.11.2026
18.11.2026
02.12.2026
16.12.2026
30.12.2026
13.01.2027
27.01.2027
10.02.2027
24.02.2027
10.03.2027
To be continued

Axio Scan 7

Whole Slide Imaging Induction: Course Syllabus

System: ZEISS Axioscan 7 High-Performance Slide Scanner

I. Introduction to Automated Slide Scanning

The Axioscan Concept: Moving from manual microscopy to "Digitize & Analyze" workflows.
Imaging Modalities:
- Brightfield: For H&E and IHC-stained histology.
- Fluorescence: High-speed multiplexing (up to 9 channels) using Colibri 7 LED sources.
- Polarization: Quantitative petrography for geological thin sections or specialized clinical applications.
Hardware Capabilities:
- Capacity: Loading up to 100 slides (standard 26 x 76 mm) for 24/7 autonomous operation.
- Cameras: Axiocam 705 (color) for brightfield and Axiocam 712 (mono) for high-sensitivity fluorescence.

II. Sample Preparation: The Golden Rules

Slide Integrity: Slides must be clean, free of excess mounting medium, and have no overhanging coverslips.
Labeling: Best practices for barcodes/QR codes to enable LIMS integration and automated metadata population.
Tray Loading: Correct orientation of slides in the magazine (up to 25 trays of 4 slides) and ensuring trays are securely seated.

III. System Startup & Software Setup

Hardware Initialization:
1. Main Power Supply.
2. Workstation Boot-up.
3. Light Sources (Colibri 7 / X-Cite).
ZEN Slidescan Interface:
1. Navigating the Magazine View for an overview of all loaded slides.
2. Scan Profiles: Using "Master" profiles vs. creating custom templates for different tissue types.
3. Storage Location: Strict policy of saving to the local D: Drive first to prevent network bottlenecks during high-speed acquisition.

IV. The Automated Workflow (Step-by-Step)

Step 1: The Preview (Pre-scan): Rapid low-magnification overview to locate tissues.
Step 2: Tissue Detection: Utilizing the Tissue Detection Wizard (or AI-based detection) to automatically define Regions of Interest (ROIs).
Step 3: Focus Strategy:
- Setting up focus maps (global vs. local focus points).
- Autofocus optimization for varied tissue thicknesses.
Step 4: Acquisition Settings:
- Brightfield: White balance and shading correction (flat-fielding).
- Fluorescence: Setting exposure times and "Fast Switching" filter wheel configurations.
- Z-Stacks/EDF: Extended Depth of Field for thick samples.

V. Post-Processing & Data Management

The .CZI Ecosystem: Navigating large multi-gigabyte files in ZEN Lite or specialized viewers (e.g., QuPath).
Stitching & Calibration: Automatic alignment of tiles and system self-calibration for reproducible image quality.
Data Export: Moving data from local scratch disks to facility servers or cloud storage platforms.

VI. Shutdown & Facility Etiquette

Safe Unloading: Using the "Unload" command before opening the magazine door to prevent mechanical jams.
Cleanup: Checking trays for broken glass or leaking oil/medium.
Shutdown Sequence: Cooling down LED sources and following the designated "Last User" shutdown procedure.
Error Reporting: How to log issues in the facility management software.

The next dates for the on-site appointments are planned from 9:00 -12:00 on:

25.02.2026
04.03.2026
25.03.2026
08.04.2026
29.06.2026
13.05.2026
27.05.2026
10.06.2026
24.06.2026
08.07.2026
22.07.2026
05.08.2026
19.08.2026
02.09.2026
16.09.2026
30.09.2026
14.10.2026
28.10.2026
11.11.2026
25.11.2026
09.12.2026
23.12.2026
06.01.2027
20.01.2027
03.02.2027
17.02.2027
03.03.2027
To be continued

Lightsheet 7

Light Sheet Fluorescence Microscopy (LSFM) Induction: Course Syllabus

System: ZEISS Lightsheet 7 (Latest 2026 Configuration)

When imaging with the Lightsheet 7, 50% of your success depends on sample mounting. If the sample moves or the Refractive Index is mismatched, the data will be unusable regardless of the software settings."

I. Foundations of Light Sheet Microscopy

The LSFM Principle: Decoupling the illumination and detection axes to achieve 90° orthogonal imaging.
Advantages of Light Sheet:
- Minimal Phototoxicity: Low light dose for long-term live-cell imaging (embryogenesis, organoids).
- High Speed: Megapixel-per-second acquisition using sCMOS cameras.
Dual-Sided Illumination: Understanding how the system uses two light sheets to compensate for absorption and scattering in large samples.
Pivot Scanning: Using the laser "pivot" to eliminate shadows/stripes caused by dense structures in the sample.

II. Sample Mounting & Optics (The Most Critical Step)

The Sample Chamber: Proper cleaning and assembly of the imaging chamber (trans-illumination vs. fluorescence).
Refractive Index (RI) Matching:
- Choosing the correct objective for the medium (Water, Clearing agents like Scale, BABB, or SeeDB).
- Adjusting the Correction Collar for specific RIs (ranging from 1.33 to 1.58).
Mounting Techniques:
- FEP Tubing: For live embryos or organoids.
- Agarose Embedding: Suspending samples in glass capillaries.
- Glue/Hooks: For large, cleared whole organs (e.g., mouse brain/kidney).

III. System Start-up & Hardware Configuration

Power-up Sequence: Components, PC, and Laser modules.
Objective Installation: Careful handling of the specialized LSFM detection and illumination optics.
Alignment (Adjust Tool):
- Calibrating the overlap of the two light sheets.
- Centering the light sheet in the focal plane of the detection objective.

IV. ZEN Software: Multi-View Acquisition

The Workflow: "Locate" for sample positioning and "Acquisition" for experiment setup.
Multi-View Setup:
- Rotating the sample (360°) to find the best imaging angle.
- Defining multiple views for later 3D reconstruction/fusion.
Advanced Modules:
- Multiside Imaging: Balancing the left and right illumination intensity.
- Z-Stacks: Setting the start/stop and "Optimal" step size for LSFM.
- Time Lapse: Managing focus drift and environmental controls (heating/CO₂).

V. Data Management (The "Big Data" Challenge)

File Sizes: A single experiment can easily exceed 500GB; discussion of the .CZI and BigDataViewer formats.
Data Streaming: Best practices for writing directly to high-speed SSD arrays (D: Drive) to avoid frame drops.
Offline Processing: Introduction to Multiview Reconstruction, Deconvolution, and Image Stitching requirements.

VI. System Maintenance & Safety

Chamber Decontamination: Strict protocols for cleaning after using toxic clearing agents or live biological samples.
Objective Care: Cleaning the specialized dipping lenses to prevent RI-matching fluid residue buildup.
Laser Safety: Procedures for a system with an open sample chamber.
Reporting: Logging usage hours and reporting any leaks or mechanical irregularities in the capillary drive.

Please contact us: Advanced training for Lightsheet 7

We will get in touch with you for a training appointment.