Core Facilities
USF Health - College of Medicine


    Leica TCS SP2 laser scanning confocal microscope

 

Confocal imaging optically reduces signals from the out-of-focus plane, thereby improving image contrast and resolution. The Leica SP2 system consists of a Leica inverted microscope equipped with motorized z-stage and multiple laser lights, which allows optical sectioning of multiply labeled fixed and live cells. Four laser sources cover essentially the entire range of visible light spectrum, including Diode Violet/Blue laser (405nm), Ar (458, 476, 488, 514 nm), Yellow DPSS (561 nm) and HeNe (633nm). Any dye or fluorophore that is excited by one of these wavelengths can be used.  Using unique prizm-based light separation and spectral unmixing, SP2 allows fine-tuning of the range for collecting emission light, thus permitting image acquisition of a variety of dyes.  It also offers tools for fluorsecence recovery after photobleaching (FRAP) and fluorescence resonance energy transfer (FRET) to study dynamics of macromolecules and organelles and a various programs for analyzing images including simple 3D construction and quantification.  The inverted microscope allows use of the open cell culture chamber (Harvard Apparatus PDMI-2) and microinjection and micromanipulation systems for drug treatment and cell manipulation during live cell imaging.

Leica SP2 Manual is available for download and view for first time users.

Features

  • Point-laser scanning fluorescence microscopy of fixed and live samples with UV and visible lasers and phase contrast imaging.
  • Spectral prism and slit for precise spectral imaging of various dyes and fluorescent proteins.
  • Z-series for 3D image.
  • Time series for time-lapse imaging.
  • FRAP (fluorescence recovery after photobleaching), FRET (fluorescence resonance energy transfer) and photoactivation.

Specification

  • Leica DMIRE2 inverted microscope with motorized z-stage (z-Galvo and z-Wide).
  • Objective lenses
    • 10x/ 0.40 PLAN APO
    • 20x/ 0.70 PLAN APO
    • 40x/ 1.25-0.75 PLAN APO Oil
    • 63x/1.40-0.60 PLAN APO Oil
    • 63x/1.20 PLAN APO Water
    • 100x/1.40-0.70 PLAN APO Oil
  • Fluorescent filters
    • A for 340-380 nm excitation and viewing of DAPI, Hoechst
    • I3 for 450-490 nm excitation and viewing of greenish fluorophores, such as YFP, GFP, Alexa-488, FITC.
    • N2.1 for 513-560 nm excitation for redish fluorophores, like Cy-3, TRITC, DsRED.

 Excitation Lasers

Laser source

Excitation wavelength

Dyes & Fluorophores

Emission color

405 Diode laser

405 nm

DAPI, Hoechst

Blue

Argon laser

458 nm

CFP

Cyan

476 nm

 

Cyan-Green

488 nm

Alexa 488, Oregon Green, FITC, GFP,

EGFP, DiO, Cy2

Green

514 nm

YFP, EYFP

Yellow

Red HeNe laser

633 nm

Alexa 633, Alexa 647, Cy5, TO-PRO3,

Far red


Applications

Colocalization
FRAP
FRET