Wright Cell Imaging Facility

Upright Confocal   Upright Widefield   Live Cell Inverted    Microdissection   Stereology   Offline Analysis

Zeiss LSM 510  META

Confocal microscopes acquire 'optical sections' of fluorescent specimens. This is achieved by placing a pinhole in front of the emitted-light detector. Only light from the focal point reaches the detector; light from above and below the focal point does not get through the pinhole.

A series of such sections can be acquired in the z-axis of the specimen and reconstructed allowing 3-D visualisation and analysis.

This confocal system is equipped with a META-detection system This allows discrimination of similar coloured fluorophores (e.g. GFP, YFP) that cannot be separated by traditional filtering. It may also be useful in separating green fluorophores from green-yellow autofluorescence.

Aside from the more typical visible-light lasers, our Zeiss 510 also has an ultra-violet laser for short wavelength dyes such as DAPI and Hoechst.

3D-reconstruction of  DAPI (blue)-stained nuclei in a rat spinal cord section. Intra-ventricular DiI injection results in red-staining of cells in the ependymal cell layer of the central cord. Image provided by A. Mothe (TWRI). Scale-bar 10 µm. Image processed with ImageJ running VolumeJ.

Zeiss 510 laser lines

Laser

Line (nm)

Example dyes

Argon

351, 364

DAPI, Hoechst

Argon

458, 477, 488

FITC, GFP

Green HeNe

543

Cy3, DsRed

Red HeNe

633

Cy5

Objectives on the LSM510

5× FLUAR, 0.25 NA
10× FLUAR, 0.5 NA
25× PLAN NEOFLUAR DIC, 0.8 NA, Water imm
63× PLAN APOCHROMATIC, 1.2 NA, Water imm.
63× ACROPLAN, 0.95 NA, Water (dipping).
63× PLAN-APO, 1.4 NA Oil imm.

08/02/2006