Confocal LSM 510

 

Introduction

In confocal microscopy, placement of a variable spatial pinhole at the confocal plane of the lens eliminates out of focus light thus reducing background interference.

The LSM 510 meta is a Zeiss laser scanning confocal system attached to an inverted motorized microscope (Axiovert 200M). It has three PMTs and the Meta detector which allows simultaneous acquisition of up to 8 channels of the emission spectrum to separate fluorophores with overlapping emission spectra or as an additional PMT.

 

Laser line Wavelength Power Suitable fluorophores
Multiline Argon 458, 477, 488, 514 30mW Cy2, GFP, Mitotracker Green, JC1
Diode-pumped solid-state 561 10mW Cy3, TAMRA, PI, Texas Red, PKH26
Helium-Neon 633 5mW Cy5, APC, Alexa 647, TO-PRO-3Cy2, GFP, Mitotracker Green, JC1
UV Diode 405 30mW DAPI, Hoechst

 

Objective lens Magnification NA coverslip Working distance Immersion
EC Plan Neofluar x5 0.15 0.17 mm 18.5 mm Air
EC Plan Neofluar x10 0.3 5.2 mm Air
LCI Plan Neofluar x25 0.8 0-0.17 mm 0.21-0.17 mm Oil Glycerin Water
EC Plan Neofluar x40 1.3 0.17 mm 0.21 mm Oil
EC Plan Apochromat x63 1.4 0.17 mm 0.19 mm Oil
C Apochromat x40 1.2 0.14-0.19 mm 0.28 mm @ cover glass 0.17 Water

 Acquisition

Acquisition is performed with the Zeiss proprietary ZEN 2009 software.
Three PMT detectors and the META detector for simultaneous detection and one transmitted-light PMT allow superimposing fluorescence images on brightfield or DIC images.
The software allows 360° XY scanning field rotation, and freely definable regions of interest (ROI).
Acquisition modes: Spot, line/spline, frame, lambda stack.
Resolution up to 2048 x 2048 pixels, scanning speed up to 5 frames/sec at 512 x 512 pixels.

Special applications

Photobleaching, live imaging, ROI, positions, tiling, time series and combination (XYZ|T), increased intensity over Z.

Analysis 

The Zen software provides a wide range of 2D/3D image processing tools: export to TIFF, maximum projection, 3D reconstruction, 2D colocalization, spectral unmixing and more. For basic visualization and processing you can download the free ZEN lite edition software. An offline full version is available for analysis at Computer Analysis 4. Output files can be opened by Imaris and ImageJ/FIJI for analysis.

Instructions for specific applications

New users

Please contact Edith Suss-Toby, tel. +972-4-8295347/61 to coordinate a meeting.
Tips for immunofluorescence experiments.
Please prepare controls before starting a new model.

Reading materials prior to the first instruction session:

Reservations

The system can be reserved in the BookItLab online reservation system, Confocal LSM 510 Meta Inverted microscope. For live experiments, the heating stage insert should be reserved separately.

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