LTSEM image of a diatom biofilm LTSEM cross section of a biofilm

Low Temperature Scanning Electron Microscope (L.T.S.E.M.)

Low Temperature Scanning Electron Microscopy (LTSEM) is a high resolution imaging tool used in the SERG laboratory to examine soft (usually muddy sediment) samples, with minimum disruption to structure integrity and without the addition of chemicals.

The samples are frozen in situ with Liquid Nitrogen [ LN2] (which has a temperature of -196C) by lifting a small amount of surface material on a thin aluminium strip(surface scrape) and plunge freezing it. This is used for studying the surface of the sediment. The other method involves using a cryolander ( a specialist freezing tool developed in SERG by Blackburn and Wiltshire) which sits on the surface of the sediment (without disturbing the sample sediment) through which LN2 is poured until the sediment is frozen to a depth of 1cm. This is used mainly for depth profiling and structural studies. This allows for comparisons between laboratory work and scanning electron images e.g. water content and structure or chlorophyll content and diatom depth , carbohydrate content and E.P.S (extra-polymeric-substance) content; visualising the binding of diatoms, sand grains and other sediment structures. Samples are then stored in LN2, or a freezer, for later examination under the LTSEM.

The Scanning Electron Microscope produces electrons from a tungsten filament under a high voltage. These electrons are accelerated via an anode through a series of electromagnetic lenses and then through scanning coils above an objective aperture which scan the electron beam over the surface of a specimen. The resulting energies (electrons, light, x-rays) can then be collected and visualised or quantified. The SERG Jeol 35CF scanning electron microscope collects secondary electrons which give a (pseudo) three dimensional view of the specimen with a large depth of focus (millimetres) this combined with very high resolution makes the scanning electron microscope ideal for examining sediments. The samples are manipulated into small holders under LN2 where they are subsequently transferred into the microscope; where they can be partially freeze dried within the controlled environment of the LTSEM The sample can be observed during freeze drying until enough extraneous water has come off to allow a clear view of the specimen. The sample is then gold sputter coated (still at LN2 temperatures) to maximise visualisation while minimising energy build-up.