Laser Scanning Confocal Microscopy (LSCM)

Technique in brief

The confocal is a microscope which focusses only on a single focal plane. Lasers are used for point illumination and the resulting fluorescence is detected by a photodetector. The design of a confocal ensures that only light produced by fluorescence close to the focal plane is observed. By scanning across the sample line by line an image may be produced.

One of the advantages of a confocal microscope over a normal one is the ability to shift focal planes. Successive images generated at different focal plans can be combined to produce 3D images.

Current instrumentation

The current LSCM at the AAC is a Zeiss 710 with and inverted sytle microscope stage and 5 lasers available at the following wavelengths:

• 405nm

• 488nm

• 514nm

• 561nm

• 633nm

Applications

Confocal microscopy is used mainly but not exclusively for biological and life science applications. In general a variety of specific fluorescent stains or fluorphores are applied to a thin sections of biological material however other opaque and reflective materials can be observed given the right conditions.

Sample requirements

In general sample must be prepared for an inverted microscope, for biological samples a sealed cover slip must be used.

Considerations should include: Coverslip thickness and spacers, the opaque nature of the sample, sample thickness, mounting media and fluorescent probe application.

For further information

Contacts: Shane Askew