Leica SP8 with whitelight laser inverted confocal

RESERVE THIS MICROSCOPE

Leica SP8 with whitelight laser inverted confocal

Confocal imaging using a super continuum, white light (WLL) pulsed laser source that allows users to specify any excitation wavelength between 470nm and 670nm. Together with an AOBS (acoustical optical beam splitter) up to 8 discrete laser lines can be selected simultaneously with a 1 nm precision. Because the WLL system is a pulsed laser it can be used in conjunction with hybrid detectors to provide gated suppression (removal) of autofluorescence and reflected light. The SP8's spectral detection system uses a prism spectrometer based emission separation (not filters) that allows user selection of emission detection bandwidth as well as spectral scanning of emission profiles. This allows users to precisely select any emission bandpass they require. Additionally, the system can scan at 8000Hz for ultra-fast scanning (and greatly reduced photobleaching) via its selectable resonant scanner. It is fully enclosed for heating and environmental control with media perfusion capabilities and CO2 control. The system is ideal for imaging live cells, tissue in culture dishes and chambered coverglass (#1.5), with AFC adaptive focus control for focus maintenance, Navigator module for tiling and Lightning module for deconvolution.

SP8 gSTED (older generation)

Stimulated Emission Depletion (gSTED) is a super resolution method. The combination of pulsed WLL and hybrid detectors has allow the development of gated or gSTED. gSTED has improved resolution (< 50 nm resolution in x,y) and contrast over continuous wave (cw) STED using significantly less laser intensity than cwSTED allowing the application of STED super resolution in both fixed and living samples. The SP8-gSTED uses a 592 nm depletion laser. More info on sample prep and STED fluorophore options can be found here.

System general specifications

DMI 6000 inverted microscope platform with motorized x,y stage with z-galvo stage

Objectives

Lasers

WLL2 Laser; avg. power 1.5 mW: 470nm to 670 nm, 78 Mhz;
with integrated pulse picker: 78, 39, 19.5, 9.75 MHz
405nm/AOTF laser 50mW
Argon laser (FRAP, bleach), 65 mW: 458, 476, 488, 496, 514 nm
STED (depletion) laser 592nm

Leica tandem galvo/resonant scanner system

Variable Scanner 1 - 1400 Hz Line frequency max: 2800 Hz 5 Images/second, 512 x 512 ~54 Images/second, 512 x 16 Max. Image resolution 8192 x 8192 Line frequency up to 2800 Lines/second. Resonant scanner, 8 kHz X2Y-Scanner for high frame rate: 28 Images/second, 512 x 512 ~290 Images/second, 512 x 16 Max. Image resolution 1024 x 1024 Line frequency up to 16000 Lines/second Very big scanfield with 12,52 mm (SFZ) diagonal in the Intermediate image plan Hardware zoom, stepless 1,3x - 48x
Scan head electronics updated from 40MHz to 80MHz. 

Detection

5 channel confocal detection: 3 Hybrid-GaAsP gated detectors for increased light sensitivity, 2 standard fluorescent photomultiplier tubes (PMT)
1 transmitted light detector with DIC/phase options

Software

Leica LAS X Premium: control software, Navigator, Lightning, Dye Finder, MicroLab, FRAP Zoomer, FRAP, FRAP-XT, FRET-AB, FRET-SE; 

Off-hour credits

Labs that use the following microscopes: Zeiss LSM880, Leica Stellaris8, and/or Leica SP8 for 25 or more hours per month, during OFF-HOURS will be rebated to reach $32/hour for all use of that microscope.
Labs that use the Zeiss LSM880, Leica Stellaris8, and/or Leica SP8, for 75 or more hours per month, during OFF-HOURS will be rebated to reach $23/hour for usage. This rate is in recognition of the high number of hours required for continuous overnight and weekend imaging inherent in the use of these microscopes for live cell imaging.
To qualify for these rates you must have after-hours access to the facility (i.e. be an experienced, competent user) and must use the confocal/2 photon between 5:00 pm and 8:00 am M-F, holidays, or any time on weekends. Reserved but unused time does not qualify for these discounts. Please speak with the facility Director Jon Mulholland for more information.

Funding

This microscope was funded by a NIH S10 Shared Instrumentation Grant Award Number 1S10OD010580 (Dr. Margaret Fuller, PI) with significant contribution from Stanford's Beckman Center. Installation date: 11/2013

Required Acknowledgement

The project described was supported, in apart, by Award Number 1S10OD010580 from the National Center for Research Resources (NCRR). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NCRR or the National Institutes of Health.

Location