Leica TCS SP8 Confocal Laser Scanning Microscope
Overview
Laser scanning confocal microscopes are designed to image thick and/or multi-colour labelled samples. The Leica TCS SP8 uses Leica’s spectral (SP) detector technology which provides the flexibility of different emission wavelengths without being restricted to equipped filters. The system uses an adaptive focus control (AFC) system which maintains focus over time to improve the stability of long-term experiments. In addition the microscope is equipped with 3 HyD super-sensitivity detectors. Those detectors perform especially well in low signal intensity conditions and for live cell imaging. The Leica HyBrid (HyD) detectors can be used in 3 different detection modes. The first mode is similar to a standard PMT confocal detector. The second mode is called “BrightR” which is useful for highly dynamic samples with large intensity spectra within the same sample. The photon counting mode gives you the opportunity of quantitative imaging. The SP8 has an optional on-stage incubator to control temperature and CO2 to create the ideal environment for live cell imaging.
Taken this together the Leica TCS SP8 is a high-end imaging platform for microscopy that requires high sensitivity and for the flexibility to detect samples with non-standard emission wavelengths. Additionally, the SP8 is capable of performing long term studies that require high focus stability.
The Bioimaging Centre houses three different Leica TCS SP8 systems (for specifications see below) which can be booked by our users.
The microscope is located within the Bioimaging Centre and is operated in a user-priority scheme with 80% of usage reserved to support research members of Sarah Gurr, Gero Steinberg and Nick Talbot. For the remaining time the system is bookable as all other Bioimaging equipment.
Advantages
- Ability to control depth of field, elimination or reduction of background information
- Capability to collect serial optical sections from thick specimens
- Imaging of multiple labels at the same time
- Flexibility to detect samples with non-standard emission wavelengths
Disadvantages
- Low scan speed
- Less sensitive then Spinning disk confocal microscope
Specifications of Bioimaging SP8
Motorised inverted microscope
Objectives
- HC PL APO 10x/0.4 Dry
- HC PL APO 20x/0.7 Dry
- HC PL APO 40x/1.3 Oil
- HC PL APO 63x/1.4 Oil
- HC PL APO 63x/1.2 Water
- HC PL Apo 100x/1.44 Oil
Laser Lines
| Laser | Excitation Lines (nm) | Colour of fluorophores | Examples of fluorophores |
|---|---|---|---|
| Blue diode | 405 | Blue | DAPI / Alexa 350 |
| Argon | 458 / 476 / 488 / 496 / 514 | Green/Cyan and Yellow | GFP / YFP / Alexa 488 / FITC |
| DPSS561 | 561 | Orange and Red | DsRed / TagRFR / Alexa 568 |
| HeNe 594 | 594 | Red | Alexa 594 / Texas red /mCherry |
| HeNe 633 | 633 | Far-red | Alexa 633 / CY5 |
- 2 conventional detectors(PMTs)
- 3 Hybrid detectors (HyD)
- On stage Incubator with continuous temperature and CO2 feedback control for live cell work
The microscope is located in the teaching lab and can be booked by all Bioimaging users out of term time as well as out of hours/weekends.
Advantages
- Ability to control depth of field, elimination or reduction of background information
- Capability to collect serial optical sections from thick specimens
- Imaging of multiple labels at the same time
- Flexibility to detect samples with non-standard emission wavelengths
Disadvantages
- Low scan speed
- Less sensitive then Spinning disk confocal microscope
Specifications of Teaching Lab SP8
Objectives:
- HCX PL Fluotar 10x/0.30 Dry
- HC PL APO 20x/0.75 IMM
- HC PL APO 40x/1.30 Oil
- HC PL APO 63x/1.40 Oil
- HCX PL APO 100x/1.44 Oil
Laser Lines
| Laser | Excitation Lines (nm) | Colour of fluorophores | Examples of fluorophores |
|---|---|---|---|
| Blue diode | 405 | Blue | DAPI / Alexa 350 |
| OPSL488 | 488 | Green | GFP / Alexa 488 / FITC |
| OPSL552 | 552 | Red | mCherry / TagRFR / Alexa 568 |
- 2 conventional detectors(PMTs)
- 3 Hybrid detectors (HyD)
- On stage Incubator with continuous temperature and CO2 feedback control for live cell work
This microscope achieves super-resolution (lateral 140 nm, axial 350 nm resolution) by combining sub-airy pinhole sizes with fast system-integrated data post-processing (10x faster deconvolution). The system combines the extremely sensitive HyD detectors with a resonance scanner for increased speed and sensitivity. An upright microscope configuration using water-dipping objectives improves diffraction-limited spatial resolution and a long working distance is especially suitable for large specimen and tissues.
This microscope will be operated in a user-priority scheme with 80% of usage reserved to support research members of Phil Ingham and Steffen Scholpp. For the remaining time the system is bookable as all other Bioimaging equipment.
Advantages
- HyVolution II system provides super-resolution down to 140 nm using system-integrated Huygens deconvolution
- High scan speed with the 8 kHz Resonance Scanner
- Simultaneous imaging of multiple labels
- Ability to control depth of field, elimination or reduction of background information
- Flexibility to detect samples with non-standard emission wavelengths ('Whte light laser')
Specifications of SP8X HyVolution II
- Scanning stage upright microscope
Objectives
- Fluotar Phase Contrast 10x/0.32 Dry
- Fluotar Phase Contrast 20x/0.40 Dry
- Apochromat Water-Dipping, Long 20x/0.5 Water
- Apochromat Water-Dipping, Long 40x/0.8 Water
- Apochromat Water-Dipping, Long 63x/0.9 Water
Laser Lines
| Laser | Excitation Lines (nm) | Colour of fluorophores | Examples of fluorophores |
|---|---|---|---|
| White Light Laser | 470-670 | Green / Cyan, Yellow, Orange and Red | GFP / YFP / Alexa 488 / FITC / DsRed / TagRFR / Alexa 568 / Alexa 594 / Texas red /mCherry / Alexa 633 / CY5 |
- 2 Hybrid detectors (HyD)
- 2 Conventional detectors (PMT)
- Resonance scanner
- Super Z Galvanometric Piezo Stage
