Leica SP8 STED 3X

This fully spectrally tunable point scanning confocal has many unique capabilities. First, it is equipped with a white light laser (WLL). This laser allows users to choose any laser excitation wavelength from 470 nm – 670 nm in 1 nm increments, which expands the range of fluorophores that can be imaged on this system. In addition to the WLL, the system also has a 405 nm diode laser and a 440 nm pulsed diode laser. For detection, it is equipped with 5 spectral detectors – 2 multi-alikali PMTs, and 3 HyDs (GaAsP Hybrid detector) with enhanced sensitivity and photon counting capabilities. High-speed imaging using the resonant scanner is also available, allowing acquisition rates of up to 28 fps (at 512x512). Available objectives include:

10x/0.3 dry

86x/1.2 water (motCorr)

93x/1.3 Glycerol (motCorr)

100x/1.4 oil

(others available to be installed upon request – notify Facility Director prior to session)

 

FLIM: Fluorescence lifetime microscopy (FLIM) is possible on this system. The combination of pulsed lasers (WLL and 440 nm laser) with photon counting HyDs allow this capability. Fluorescence lifetime imaging allows us to measure the amount of time a fluorophore of interest spends in the excited state. This is a photochemical property of the fluorophore, and can be affected by environmental changes and interactions. FLIM FRET is also possible using this technique.

STED: In addition to the confocal and lifetime imaging capabilities, the system is capable of STED (Stimulated Emission Depletion) imaging, which is an optical super-resolution technique. This technique adds a second, donut shaped laser beam to your imaging protocol to effectively turn off your fluorophores in a particular spatial area. Resolution <40 nm laterally, and ~120 nm axially are possible. Some special considerations for sample preparation are needed for STED imaging. Please discuss with the Facility Director in advance before planning your staining protocol for STED imaging.