AS Biology Core Principles Light and Electron Microscopes

Objectives State the resolution and magnification that can be achieved by a light microscope, a transmission electron microscope and a scanning electron microscope. Explain the difference between magnification and resolution. Explain the need for staining samples for use in light and electron microscopy.

Resolving Power

Resolving Power The limit of resolution of a microscope is the smallest distance between 2 points that can be seen using a microscope This is a measure of the clarity of the image A microscope with a high resolving power will allow 2 small objects which are close together to be seen as 2 distinct objects

The Light Microscope Series of lenses through which ordinary white light can be focused Optical microscopes cannot resolve 2 points closer together than about half (200nm) the wavelength of the light used (400-750nm) Resolution of the light microscope is 200nm or 0.2µm

The Light Microscope The total magnification is the eyepiece magnification multiplied by the objective magnification The maximum magnification of a light microscope is x1500 Many cells are transparent to light; some structures can only be seen if they are stained with a coloured solution like iodine

The Electron Microscope Electrons (negatively charged, very small particles) can behave as waves The wavelength of a beam of electrons is about 0.004nm The resolution of the electron microscope is about 0.2nm Electrons are ‘fired’ from an electron gun at the specimen and onto a fluorescent screen or photographic plate There are 2 types of electron microscopy - transmission and scanning Both focus an electron beam onto the specimen using electromagnets

Transmission Electron Microscope (TEM) In transmission EM the electrons pass through the specimen. Specimen needs to be extremely thin - 10nm to 100nm The specimen is stained with electron dense materials so that electrons do not pass through the most stained structures. These look darker than the surrounding tissue. TEM can magnify objects up to 500000 times. TEM has made it possible to see the details of organelles.

Transmission Electron Microscope (TEM) Cells or tissues are killed and chemically ‘fixed’ in a complicated and harsh treatment. This means only dead specimens can be viewed The images are always black and white, (false colour can be added to printed images) This treatment can result in alterations to the cell - known as artefacts

Transmission Electron Microscope (TEM) Transmission electron micrograph of a mitochondrion from a liver cell

Scanning Electron Microscope (SEM) In Scanning EM microscopes the electrons bounce off the surface of the specimen Produce images with a three-dimensional appearance Allow detailed study of surfaces

Scanning Electron Microscope (SEM)

Question Which type of microscope would be used to view each of the following? Blood flowing through capillaries in a fishes tail. The 3D shape of a bacterium. The growth of a pollen tube. The internal structure of a chloroplast. The surface of cells lining the small intestine. The structures found in the cytoplasm of a liver cell.