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leica_stellaris_sted_flim [2026/09/03 17:21] bioimaging [Filterset (for eyepieces)] |
leica_stellaris_sted_flim [2026/09/10 15:31] (current) bioimaging [LASERs] |
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| ===== Microscope overview ===== | ===== Microscope overview ===== | ||
| - | The Leica STELLARIS STED FLIM is a [[https://www.youtube.com/watch?v=rTkjQ6Q0p5I|point scanning confocal microscope]] designed for advanced fluorescence imaging, super-resolution microscopy and fluorescence lifetime measurements. Built on the [[https://www.leica-microsystems.com/products/confocal-microscopes/p/leica-stellaris/|Leica STELLARIS platform]], it combines a [[https://www.leica-microsystems.com/science-lab/life-science/stellaris-white-light-lasers/|White Light LASER]] (WLL) excitation source with highly sensitive Power HyD detectors and prism-based spectral detection, providing flexible excitation from 440 to 790 nm and fluorescence detection up to 850 nm. It is also equipped with a 405 nm diode laser. The system is equipped with a motorized XY scanning stage, a SuperZ Galvo Stage for rapid Z acquisition, and an Adaptive Focus Control (AFC) closed-loop hardware autofocus system for long-term stability during time-lapse experiments. A large environmental enclosure with temperature and CO2 control enables live-cell imaging under physiological conditions. The confocal scanhead incorporates a field-of-view scanner with galvanometric mirrors optimized for high resolution and sensitivity. Detection is provided by multiple Power HyD detectors, including Power HyD X detector optimized for fluorescence lifetime applications, fluorescence correlation spectroscopy (FCS), and Tau-STED measurements. For super-resolution imaging, the system is equipped with STED technology, including pulsed 589 nm and 775 nm depletion lasers, enabling imaging with resolution well below the diffraction limit. The 3D STED module provides tunable super-resolution in X, Y and Z, while TauSTED Xtend combines STED and fluorescence lifetime information to improve image quality, contrast and resolution at lower light doses, making it particularly suitable for live-cell super-resolution imaging. The system supports multicolour STED imaging across much of the visible spectrum and facilitates the investigation of molecular organization, interactions and nanoscale cellular structures. The microscope also includes STELLARIS FALCON (FAst Lifetime CONtrast), providing fluorescence lifetime imaging microscopy (FLIM) at confocal imaging speeds. Fully integrated within LAS X software, FALCON supports exponential fitting, FLIM phasor analysis, pattern fitting and FLIM-FRET analysis, enabling quantitative studies of molecular interactions, environmental sensing and dynamic cellular processes. | + | {{ :sted_principle.png?240}} The Leica STELLARIS STED FLIM is a [[https://www.youtube.com/watch?v=rTkjQ6Q0p5I|point scanning confocal microscope]] designed for advanced fluorescence imaging, [[https://www.leica-microsystems.com/science-lab/life-science/a-guide-to-fluorescence-lifetime-imaging-microscopy-flim/|fluorescence lifetime imaging (FLIM)]] and super-resolution microscopy. Built on the [[https://www.leica-microsystems.com/products/confocal-microscopes/p/leica-stellaris/|Leica STELLARIS platform]], it features a [[https://www.leica-microsystems.com/science-lab/life-science/stellaris-white-light-lasers/|White Light LASER]] (440-790 nm) and 405 nm laser as excitation sources, AOBS beam splitting, highly sensitive [[https://www.leica-microsystems.com/science-lab/life-science/power-hyd-detectors/|Power HyD detectors]] and spectral detection for flexible multicolor imaging, including near-infrared fluorophores up to 850 nm, such as Cy7. The system is optimized for live-cell imaging, featuring a motorized stage, SuperZ Galvo Stage for rapid Z acquisition, hardware autofocus (AFC) and a temperature- and CO₂-controlled Okolab dark enclosure for live-cell imaging in physiological conditions. FLIM capabilities are provided by the Power HyD X detector and [[https://www.leica-microsystems.com/products/confocal-microscopes/p/stellaris-8-falcon/|STELLARIS FALCON (FAst Lifetime CONtrast)]], enabling fast lifetime imaging, [[https://www.leica-microsystems.com/science-lab/life-science/what-is-fret-with-flim/|FLIM-FRET experiments]], [[https://www.leica-microsystems.com/science-lab/life-science/phasor-analysis-for-flim-fluorescence-lifetime-imaging-microscopy/|phasor analysis]] and [[https://www.leica-microsystems.com/science-lab/life-science/tausense-technology-imaging-tools/|TauSense]] lifetime-based contrast and multiplexing tools. |
| + | For super-resolution imaging, the system is equipped with [[https://www.leica-microsystems.com/science-lab/life-science/a-guide-to-super-resolution/|2D and 3D STED technology]] using pulsed 589 nm and 775 nm depletion lasers, providing resolution well below the diffraction limit. [[https://www.leica-microsystems.com/science-lab/life-science/extended-live-cell-imaging-at-nanoscale-resolution/|TauSTED Xtend]] combines STED and lifetime information to improve image quality and resolution while reducing light exposure, making the system particularly suitable for live-cell nanoscale imaging. | ||
| * **Microscope**: [[https://www.leica-microsystems.com/products/light-microscopes/p/dmi8/|Leica DMi8 CS inverted]] | * **Microscope**: [[https://www.leica-microsystems.com/products/light-microscopes/p/dmi8/|Leica DMi8 CS inverted]] | ||
| * **LED Illumination**: [[https://www.coolled.com/products/pe-300white/|CoolLED pE-300white]] | * **LED Illumination**: [[https://www.coolled.com/products/pe-300white/|CoolLED pE-300white]] | ||
| - | * **Detectors**: [[https://www.leica-microsystems.com/science-lab/life-science/power-hyd-detectors/|Power hybrid detectors]] (Power HyD S, Power HyD X, and Power HyD R) | + | * **Detectors**: [[https://www.leica-microsystems.com/science-lab/life-science/power-hyd-detectors/|Power hybrid detectors]] |
| + | * Power HyD S | ||
| + | * Power HyD X for FLIM imaging | ||
| + | * Power HyD R for near-infrared imaging | ||
| * **Features** | * **Features** | ||
| * | [[https://www.leica-microsystems.com/science-lab/life-science/a-guide-to-super-resolution/|2D and 3D Super Resolution STED]] | * | [[https://www.leica-microsystems.com/science-lab/life-science/a-guide-to-super-resolution/|2D and 3D Super Resolution STED]] | ||
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| {{:warning.png?25|}} Data files older than **1 month** will be automatically deleted on this system, please copy your data to the GIMM server using the desktop link. | {{:warning.png?25|}} Data files older than **1 month** will be automatically deleted on this system, please copy your data to the GIMM server using the desktop link. | ||
| + | ===== Additional information for sample preparation ===== | ||
| + | * [[https://downloads.leica-microsystems.com/STELLARIS%208%20STED/Application%20Note/STELLARIS%20STED%20Sample%20Preparation%20Guide.pdf|The Guide to STED Sample Preparation (application report)]] | ||
| ===== System components ===== | ===== System components ===== | ||
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| ^ LASER ^ Wavelength ^ | ^ LASER ^ Wavelength ^ | ||
| | Violet | <fc #a600ff>405 nm</fc> | | | Violet | <fc #a600ff>405 nm</fc> | | ||
| - | | WLL | <fc #0000ff>440 nm</fc> - <fc #be0000>790 nm</fc> | | + | | WLL | <fc #0071ff>440 nm</fc> - <fc #be0000>790 nm</fc> | |
| ==== Filterset (for eyepieces) ==== | ==== Filterset (for eyepieces) ==== | ||
| Line 71: | Line 77: | ||
| ^ Magnification ^ Model ^ Immersion ^ NA ^ WD (mm) ^ Reference ^ | ^ Magnification ^ Model ^ Immersion ^ NA ^ WD (mm) ^ Reference ^ | ||
| - | | <fc #df2b37>5x</fc> | EC Plan-NeoFluar | Air | 0.16 | 13.6 | [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420330-9901-000/Objective-EC-Plan-Neofluar-5x-0.16-M27|420330-9901-000]] | | + | | <fc #fef042>10x</fc> | HC PL APO CS2 | Air | 0.40 | 2.56 | [[https://www.leica-microsystems.com/objectivefinder/detail/objective/506424/|11506424]] | |
| - | | <fc #0aa05c>20x</fc> | Plan-Apochromat | Air | 0.80 | 0.55 | [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420650-9903-000/Objective-Plan-Apochromat-20x-0.8-M27|420650-9903-000]] | | + | | <fc #0aa05c>20x</fc> | HC PL APO IMM CORR CS2 | Water | 0.75 | 0.66 | [[https://www.leica-microsystems.com/objectivefinder/detail/objective/506343/|11506343]] | |
| - | | <fc #01b3f5>40x</fc> | LD LCI Plan-Apochromat Corr | Water | 1.20 | 0.41 | [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420862-9970-799/Objective-LD-LCI-Plan-Apochromat-40x-1.2-Imm-Corr-DIC-M27|420862-9970-799]] | | + | | <fc #01b3f5>40x</fc> | HC PL APO Oil CS2 | Oil | 1.30 | 0.17 | [[https://www.leica-microsystems.com/objectivefinder/detail/objective/506428/|11506428]] | |
| - | | <fc #2c80c0>63x</fc> | Plan-Apochromat DIC | Oil | 1.40 | 0.19 | [[https://www.micro-shop.zeiss.com/en/de/shop/objectives/420782-9900-799/Objective-Plan-Apochromat-63x-1.4-Oil-DIC-M27|420782-9900-799]] | | + | | <fc #2c80c0>63x</fc> | HC PL APO Oil CS2 | Oil | 1.40 | 0.14 | [[https://www.leica-microsystems.com/objectivefinder/detail/objective/506350/|11506350]] | |
| - | + | | 100x | HC PL APO OIL STED WHITE | Oil | 1.40 | 0.13 | [[https://www.leica-microsystems.com/objectivefinder/detail/objective/506378/|11506378]] | | |
| - | ===== How to start your session ===== | + | |
| - | + | ||
| - | If you are the **first user** of the day: | + | |
| - | * Turn off Components | + | |
| - | {{::zeiss_lsm_980_components.png?100|}} | + | |
| - | * Turn off Main Switch | + | |
| - | {{::zeiss_lsm_980_main_switch.png?100|}} | + | |
| - | * Turn on the computer | + | |
| - | * Turn on Main Switch | + | |
| - | * Turn on Components | + | |
| - | * Log into Windows with your Agendo credentials | + | |
| - | * Start **ZEN** | + | |
| - | Else: | + | |
| - | * Log into Windows with your Agendo credentials | + | |
| - | * Start **ZEN** | + | |
| - | + | ||
| - | ===== How to end your session ===== | + | |
| - | + | ||
| - | If there is another user: | + | |
| - | * Clean up immersion objectives (gently wipe with optic cleaning paper and EtOH embedded paper) | + | |
| - | * Exit the ZEN software | + | |
| - | * Log off the computer | + | |
| - | If you are the **last user** of the day: | + | |
| - | * Clean up immersion objectives (gently wipe with optic cleaning paper and EtOH embedded paper) | + | |
| - | * Exit the ZEN software | + | |
| - | * Turn off the computer | + | |
| ---- | ---- | ||
| [[resources|Back to the Equipment page]] | [[resources|Back to the Equipment page]] | ||
leica_stellaris_sted_flim.1788448877.txt.gz · Last modified: 2026/09/03 17:21 by bioimaging
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