Leica SP8 STED Super Resolution Confocal

Eckstein Medical Research Building - Room 89

The Leica 3x STED system is a super resolution microscope based on the Leica SP8 confocal microscope. It has a guaranteed resolution of 50 nm though the system has been routinely achieving 40 nm. It is equipped with multiple excitation lasers including a continuum fiber laser that can deliver up to 8 simultaneous lines of any wavelengths between 470 and 670 nm. All common fluorophores can be imaged. Alexa 532, TRITC, Alexa 594 and ATTO 647N are ideal STED fluorophores. YFP, Venus and mStrawberry are excellent STED fluorescent proteins. It has 4 detectors, two of which are high-sensitivity Hybrid detectors. It can be operated in STED or standard confocal mode. Feature highlights include:

  •  Continuum Fiber Laser: continuous excitation  tuning from 470 nm to 670 nm
  • Argon Ion Laser with 457, 477, 488 and 514 nm lines
  • 405 nm DPSS laser
  • 775 and 660 nm Depletion lasers
  • Two HyD and two PMT detectors
  • Transmitted light detector
  • Objective lenses
    • 10x dry
    • 20x dry
    • 40x oil for confocal
    • 63x oil for confocal
    • 93x glycerol for STED only
    • 100x oil for STED only

Below is an example of the dramatic increase in resolution provided by STED. The sample is 23 nm beads labeled with ATTO647N (top two images) and Alexa 594. Confocal images on the left, 775 nm STED on the right. Note the typical diffraction-limited 200 nm spots in the confocal images, and the dramatic resolution improvement in the STED images.

647N-AF594 STED

 

Useful Links

Basic Instructions

STED theory and labeling strategies 

STED Sample Prep Quick Guide

Acceptor Photobleaching FRET Guide

Sensitized Emission FRET Guide

FRAP Guide

Link to Carver College of Medicine STED article

 

STED Inventor Stefan Hell wins Nobel Prize!

http://www.the-scientist.com/?articles.view/articleNo/41169/title/Nanoscopy-Wins-Nobel/