Microscope PDF

MICROSCOPE Ass. Lec . Ghufran S. Salih What is microscope? It is a device which deals with organisms too small they cannot be seen distinctly with the unaided eye . - The human eye can resolve objects of the order of 0.1 mm, while the light microscope can resolve objects on the order of 0.2 μm (20...

MICROSCOPE Ass. Lec . Ghufran S. Salih What is microscope? It is a device which deals with organisms too small they cannot be seen distinctly with the unaided eye . - The human eye can resolve objects of the order of 0.1 mm, while the light microscope can resolve objects on the order of 0.2 μm (200 nm) . The transmission electron microscope can resolve objects on the order of 0.1nm . - For example, their typical linear dimensions are: Animal cell (20–30 μm), a red blood cell (7.6 μm), a mitochondrion (2–5 μm), a nucleus (10 μm), microvilli (1μm), a cell membrane (10 nm), a microfilament (8–10 nm), a bacterium (0.5–5 μm) and a virus (10–100 nm) Microscope Types 1. Compound Microscope It is a light illuminated. The image in two dimensional. You can view individual cells, even living ones. It has high magnification. However, it has a low resolution. 2. Dissecting Microscope It is light illuminated. The image that appears is three dimensional. It is used for dissection ‫ﺗﺸﺮﻳﺢ‬to get a better look at the larger specimen. You cannot see individual cells because it has a low magnification. (also called stereo microscope‫)ﻣﺠﻬﺮ ﺗﺸﺮﻳﺤﻲ‬. 3. Scanning Electron Microscope (SEM): SEM use electron illumination. The image is seen in 3-D. It has high magnification and high resolution. The specimen is coated in gold and the electrons bounce off to give you and exterior view of the specimen. 4. Transmission Electron Microscope (TEM): TEM is electron illuminated. This gives a 2-D view. Thin slices of specimen are obtained. The electron beams pass through this. It has high magnification and high resolution. Compound Microscope Anatomy of a compound Microscope 1. lamp (light source) 2. Body tube 3.Condenser: Used to vary the intensity of light. 4. Stage and microscope slide with specimen 5. Diaphram:‫ اﻟﻐﺸﺎء‬Modulates the amount of light that passes through the slide. 6. Objective lenses (usually 10X, 40X, and 100X) on revolving nosepiece‫ﻗﻄﻌﺔ اﻟﺪوارة‬ 7. Ocular lens (usually 10X) The principle of the Light Microscope working : In order to know how the microscope works we must first have a basic understanding about how lenses bend and focus light to form images The measure of how greatly a substance slows the velocity of light is called the refractive index When light energy passes from one medium to another (i.e. AIR and GLASS) the light rays are bent at the point of interface. This process is called refraction. As parallel rays of light encounter ‫ﺗﺼﺎدف‬a convex lens, they are slowed ‫ﺗﺘﺒﺎﻃﺄ‬and bent ‫ﺗﻨﺤﻨﻲ‬towards the normal path‫اﻟﻤﺴﺎر اﻟﻄﺒﻴﻌﻲ‬. The point at which these rays converge‫ ﺗﺘﻼﻗﻰ‬is called the focal point‫اﻟﻨﻘﻄﺔ اﻟﻤﺤﻮرﻳﺔ‬. The distance between the center of the lens and the focal point is called the focal length‫ اﻟﺒﻌﺪ اﻟﺒﺆري‬. The strength of a lens is directly related to its focal length. A lens with a short focal length has a greater capacity for magnification than a lens with a longer focal length. Bright-field Microscope Called the ordinary microscope because it forms a dark image against a brighter background. Three factors determine the quality of an optical image: a. Magnification • A compound microscope uses two sets of lenses, with differing focal lengths, to facilitate magnification. Total magnification is calculated by multiplying the objective and eyepiece magnification together. Magnification (total) = magnification (obj. lens) x magnification (ocu. lens) Example: Mag (obj) = 40X and Mag (ocular) = 10X Mag (total) = (40X) (10X) = 400X. b. Resolution It is the ability to observe two nearby objects as distinct objects The resolution increased by: 1- using immersion oil that leads to reduce the refraction of light rays(through the air in the space between the specimen and the objective lens.) and increasing the amount of light that enters the objective lens . Only the highest power (100X) objective on the microscope is designed for use with immersion oil. 2- decreasing the wavelength of light that is used to illuminate the specimen (use blue light instead of white light). c. Contrast Contrast is the ability to detect different regions of the specimen on the basis of intensity or color. Microbes are composed of water, nucleic acids, proteins, and lipids. Most appear colorless against a colorless background when observed using bright field microscopy. Therefore in order to see them, we must devise a way to increase the contrast by: 1- Direct staining of the microorganisms 2- Indirect (negative) staining of the background. In order to stain a specimen, it must first be fixed to the slide and chemically altered. This results in the death of a specimen Microscope Care • Always carry with 2 hands • Never touch the lenses with your fingers. • Only use lens paper for cleaning • When you are finished, rotate the nosepiece to low power objective and roll the stage down to lowest level