‫المرحلة الرابعة‬ ‫ قسم تقنيات المختبرات الطبية‬/ ‫كلية المأمون‬

‫ كمال عوني‬.‫ د‬:‫استاذ المادة‬ ‫ االمراض النسجية‬: ‫المادة‬

Electron Microscope (EM)

Electron microscopy (EM) is a technique for obtaining high

resolution images of biological and non-biological specimens. It is used
in biomedical research to investigate in detailed structure of tissues, cells,
organelles, macromolecular complexes, biopsy samples, metals, and
crystals. The high resolution of EM images results from the use of
electrons (which have very short wavelengths) as the source of
illuminating radiation.

Electron microscopy is used in conjunction with a variety of

ancillary techniques (e.g. thin sectioning, immuno-labeling, negative
staining) to answer specific questions. EM images provide key
information on the structural basis of cell function and of cell disease.

An EM has greater resolving power than a light microscope and

can reveal the structure of smaller objects because electrons have
wavelengths about 100,000 times shorter than visible light photons. The
electron microscope uses electrostatic and electromagnetic lenses to
control the electron beam and focus it to form an image. These electron
optical lenses are analogous to the glass lenses of a light optical
microscope.

How electron microscopes work

If you've ever used an ordinary microscope, you'll know the basic idea
is simple. There's a light at the bottom that shines upward through a thin
slice of the specimen. You look through an eyepiece and a
powerful lens to see a considerably magnified image of the specimen
(typically 10–200 times bigger). So there are essentially four important
parts to an ordinary (light) microscope:
 ‫المرحلة الرابعة‬ ‫ قسم تقنيات المختبرات الطبية‬/ ‫كلية المأمون‬
‫ كمال عوني‬.‫ د‬:‫استاذ المادة‬ ‫ االمراض النسجية‬: ‫المادة‬

1. The source of light.

2. The specimen.
3. The lenses that makes the specimen seem bigger.
4. The magnified image of the specimen that you see.

In an electron microscope, these four things are slightly different.

1. The light source is replaced by a beam of very fast moving

electrons.
2. The specimen usually has to be specially prepared and held inside a
vacuum chamber from which the air has been pumped out (because
electrons do not travel very far in air).
3. The lenses are replaced by a series of coil-
shaped electromagnets through which the electron beam travels. In
an ordinary microscope, the glass lenses bend (or refract) the light
beams passing through them to produce magnification. In an
electron microscope, the coils bend the electron beams the same
way.
4. The image is formed as a photograph (called an electron
micrograph) or as an image on a TV screen.

Types of Electron Microscope (EM):

1) Transmission electron microscope TEM:

2) Scanning electron microscope SEM:
3) Reflection electron microscope REM.
4) Scanning transmission electron microscope STEM.

The transmission electron microscope is used to view thin

specimens (tissue sections, molecules, etc) through which electrons
can pass generating a projection image. The TEM is analogous in
 ‫المرحلة الرابعة‬ ‫ قسم تقنيات المختبرات الطبية‬/ ‫كلية المأمون‬
‫ كمال عوني‬.‫ د‬:‫استاذ المادة‬ ‫ االمراض النسجية‬: ‫المادة‬

many ways to the conventional (compound) light

microscope. TEM is used, among other things, to image the
interior of cells (in thin sections), the structure of protein molecules
(contrasted by metal shadowing), the organization of molecules in
viruses and cytoskeletal filaments (prepared by the negative
staining technique), and the arrangement of protein molecules in
cell membranes (by freeze-fracture).
Conventional scanning electron microscopy depends on
the emission of secondary electrons from the surface of a
specimen. Because of its great depth of focus, a scanning electron
microscope is the EM analog of a stereo light microscope. It
provides detailed images of the surfaces of cells and whole
organisms that are not possible by TEM. It can also be used for
particle counting and size determination, and for process
control. It is termed a scanning electron microscope because the
image is formed by scanning a focused electron beam onto the
surface of the specimen in a raster pattern. Appropriately equipped
SEMs (with secondary, backscatter and X-ray detectors) can be
used to study the topography and atomic composition of
specimens, and also, for example, the surface distribution of
immuno-labels.

Sample preparation:

Materials to be viewed under an electron microscope may require

processing to produce a suitable sample. The technique required varies
depending on the specimen and the analysis required:

Chemical fixation: For biological specimens aims to stabilize the

specimen's mobile macromolecular structure by chemical cross linking of
proteins with aldehydes such as formaldehyde.
 ‫المرحلة الرابعة‬ ‫ قسم تقنيات المختبرات الطبية‬/ ‫كلية المأمون‬
‫ كمال عوني‬.‫ د‬:‫استاذ المادة‬ ‫ االمراض النسجية‬: ‫المادة‬

Cryofixation: Freezing a specimen so rapidly, to liquid nitrogen or even

liquid helium temperatures, that the water forms (non-crystalline).

Dehydration: Freeze drying, or replacement of water with organic

solvents such as ethanol or acetone.

Embedding: After dehydration, tissue for observation in the transmission

electron microscope is embedded so it can be sectioned ready for
viewing. To do this the tissue is passed through a 'transition solvent' such
as Propylene oxide.

Sectioning : Produces very thin slices of specimen, semitransparent to

electrons. These can be cut on an ultramicrotome with a diamond knife to
produce ultra-thin slices about 60–90 nm thick. Disposable glass knives
are also used because they can be made in the lab and are much cheaper.

Staining : Uses heavy metals such as lead, uranium or tungsten to scatter

imaging electrons and thus give contrast between different structures,
since many (especially biological) materials are nearly "transparent" to
electrons (weak phase objects).

Disadvantages:

1) Electron microscopes are expensive to build and maintain.

2) Its designed to achieve high resolutions must be housed in
stable buildings (sometimes underground) with special services
such as magnetic field cancelling systems.
 ‫المرحلة الرابعة‬ ‫كلية المأمون ‪ /‬قسم تقنيات المختبرات الطبية‬
‫استاذ المادة‪ :‬د‪ .‬كمال عوني‬ ‫المادة ‪ :‬االمراض النسجية‬

‫‪Figure 1: Transmission Electron Microscope.‬‬

‫‪Figure 2: scanning Electron Microscope.‬‬

 ‫المرحلة الرابعة‬ ‫كلية المأمون ‪ /‬قسم تقنيات المختبرات الطبية‬
‫استاذ المادة‪ :‬د‪ .‬كمال عوني‬ ‫المادة ‪ :‬االمراض النسجية‬

‫‪Figure 3: shows Mycobacterium tuberculosis by using Electron microscope‬‬