Transmission electron microscope

TEM principle, instrumentation, advantages, disadvantages

1. NANO SCIENCE TRANSMISSION ELECTRON MICROSCOPE BY U. Monika I- M.Sc Biochemistry

2. INTRODUCTION ELECTRON MICROSCOPE - System of electromagnetic coils where electron beams are used as a source of illumination. Electron microscope = Magnification is high. M agnification of 2000 times than that of light microscope.

3. TYPES OF ELECTRON MICROSCOPE 1. Transmission Electron Microscope (TEM) 2. Scanning Electron Microscope (SEM)

4. TRANSMISSION ELECTRON MICROSCOPE TEM is a special type of microscope that uses electron for magnification. Electrons - smaller wavelength. Achieve extreme magnification. Average TEM - Magnification of 1,000,000x

5. DEFINITION : Electron Microscope in which electron beam is passed through the specimen to produce its image. First TEM was designed by Max knoll and Ernest ruska in 1931. TEM is a powerful tool for Material science. Reveals the finest details of internal structures.

6. PRINCIPLE Basic principle of electron microscope is similar to that of the ordinary compound microscope. Electron Beam - Light Beam Electromagnetic coils - Optical lenses. When light voltage current is passed through a filament of cathode ray tube, electron beams are produced from the filament.

7. If some voltage of current is applied to electromagnetic coils kept around the path of electron beam, the direction of electron beam can be changed suitably to focus on the specimen. When an electron beam is passed through a specimen stained with metallic gold or osmium, the specimen absorbs some rays and reflects some rays to pass through it.

8. Interaction between electrons and specimen in the beam produces the image of the specimen. Image of the specimen can be collected by an objective lens (an electromagnetic coil) Magnified by an Amplifier (another coil) Due to electron distribution, image cannot be seen with naked eyes. Image - Recorded on a Screen or Camera.

9. INSTRUMENTATION TEM consists of electron gun, condenser lens, objective lens, amplifier lens, projector lens & fluorescent screen. ELECTRON GUN - Source of electron beam CONDENSER LENS - Located below the electron gun Collect and concentrate the electron into a strong beam before focusing onto specimen.

10. SPECIMEN STAGE - Below the condenser lens OBJECTIVE LENS - Another electromagnetic coil, placed below the specimen stage. It collects the images of the specimen and focuses it towards the amplifier lens. AMPLIFIER LENS - Magnifies image produced by the objective lens to several 1000 T imes. PROJECTOR LENS - Collects the magnified image & focus it onto fluorescent screen.

11. SAMPLE PREPARATION The biological samples have to be loaded with heavy atoms like gold or osmonium. SAMPLE PREPARATION AND EXAMINATION : 1 . Wet specimen is dehydrated using ethanol or acetone. 2. Fixation is done by Osmonium tetroxide, glutaraldehyde, potassium permanganate,formalin etc.

12. 3. The fixed specimen is embedded in hard embedding medium like aradilite and cut into thin section of 50-100nm thickness using ultramicrotome. 4. Thin sections subjected to metallic staining and placed on the specimen stage between the condenser lens coil and objective coil.

13. IMAGING A Transmission Electron Microscope produces a high-resolution, black and white image from the interaction that takes place between prepared samples and energetic electrons in the vacuum chamber. Air needs to be pumped out of the vacuum chamber, creating a space where electrons are able to move. The electrons then pass through multiple electromagnetic lenses.

14. These solenoids are tubes with coil wrapped around them. The beam passes through the solenoids, down the column, makes contact with the screen where the electrons are converted to light and form an image. The image can be manipulated by adjusting the voltage of the gun to accelerate or decrease the speed of electrons as well as changing the electromagnetic wavelength via the solenoids. The coils focus images onto a screen or photographic plate.

15. During transmission, the speed of electrons directly correlates to electron wavelength; the faster electrons move, the shorter wavelength and the greater the quality and detail of the image. The lighter areas of the image represent the places where a greater number of electrons were able to pass through the sample and the darker areas reflect the dense areas of the object. These differences provide information on the structure, texture, shape and size of the sample.

16. SAMPLE PROPERTIES : S amples need to have certain properties. They need to be sliced thin enough for electrons to pass through, a property known as electron transparency. Samples need to be able to withstand the vacuum chamber and often require special preparation before viewing. Types of preparation include dehydration, sputter coating of non-conductive materials, cryofixation, sectioning and staining.

17. APPLICATIONS Ideal tool for study of ultra structure of cells. Identification of plant and animal viruses based on their structural features. Employed in the localization of nucleic acid, enzymes and proteins in cell and cell organelles. Used in cancer research for the cytological observation of cancer cells.

18. Used in various fields such as nanotechnology, life sciences, medical, biological, material research, forensic analysis, metallurgy as well as in industries and education. Used in the production and manufacturing of computer and silicon chips. Provide topographical, morphological, compositional and crystalline informations.

19. Advantages A Transmission Electron Microscope is an impressive instrument with a number of advantages such as: TEMs offer the most powerful magnification, potentially over one million times or more. TEMs have a wide-range of applications and can be utilized in a variety of different scientific, educational and industrial fields

20. TEMs provide information on element and compound structure Images are high-quality and detailed TEMs are able to yield information of surface features, shape, size and structure They are easy to operate with proper training

21. Disadvantages Some limitations of electron microscopes include: TEMs are large and very expensive Laborious sample preparation Operation and analysis requires special training Samples are limited to those that are electron transparent, able to tolerate the vacuum chamber and small enough to fit in the chamber. TEMs require special housing and maintenance Images are black and white

22. What is the Cost? TEMs are manufactured by companies such as Jeol, Philips and Hitachi and are extremely expensive. Examples of prices for new TEM models include $95,000 for a Jeol and Philips and $100,000 for a Hitachi. In India, the cheapest one costs about 80-90 lakhs.

23. SUMMARY TEM is useful for small, nanoscale analytes. TEM can create 3D images of samples. TEM can be modified for different atoms and molecules. TEM is not cheap. It is Good!