microscope

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MICROSCOPE

MICROSCOPE:

Microscope is the combination of two words " micro " meaning small " scope " meaning view. Instrument for examination of cytological and histological specimens. The science of investigating small objects using such an instrument is called microscopy MICROSCOPE

History of microscope:

History of microscope Many people experimented with making microscopes The first microscope was 6 feet long!!! The Greeks & Romans used “lenses” to magnify objects over 1000 years ago. Hans & Zacharias, Holland in 1590. 1620-Christian Huygens, another Dutchman, developed a simple 2-lens ocular system that was chromatically corrected

History of microscope:

History of microscope Galileo made first microscopic observations of an insect’s eye .He described principles of both the lenses & light and then improved upon both microscope & telescope In 1665 Robert Hooke developed a first laboratory compound microscope.

History of microscope:

Antony van Leeuwenhoek (1632-1723) is generally credited with bringing the microscope to the attention of biologists. A tradesman of Delft, Holland. 1661-He discovered bacteria, sperm cells, blood cells, microscopic nematodes etc. History of microscope

Lenses:

6 Lenses Focus light rays at a specific place called the focal point Distance between center of lens and focal point is the focal length short focal length more magnification Lens

CLASSIFICATION:

CLASSIFICATION

Types of Microscopes:

Types of Microscopes

Types of Microscopes:

Types of Microscopes

TYPES OF MICROSCOPE:

TYPES OF MICROSCOPE Simple microscope Compound microscope Dark Ground Microscope Phase Contrast Microscope Polarizing Microscope Fluorescent Microscope Electron Microscope Newer Microscopes Types of Microscopes

Simple Microscope:

Simple Microscope Similar to a magnifying glass and has only one lens. Simple Microscope

Compound Microscope:

Compound Microscope Light passes through 2 lenses Most widely used Can magnify up to 2000x Compound Microscope

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The passage of light through two lenses forms the virtual image of the object seen by the eye. Principle of Compound microscope

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Produces a dark image against a brighter background Has several objective lenses Parfocal microscopes remain in focus when objectives are changed Total Magnification P roduct of the magnifications of the ocular lens and the objective lens Bright Field Microscope

Dissecting Microscope:

Used for dissection to get a better look at the larger specimen A dissection microscope is light illuminated. Image is three dimensional . Individual cells can not be seen because it has a low magnification. Dissecting Microscope Dissecting Microscope

TERMS AND DEFINITIONS:

TERMS AND DEFINITIONS Principle of Microscopy is to get a magnified image, in which structures may be resolved which could not be resolved with the help of an unaided eye. Magnification i s the ratio of the size of an object seen under microscope to the actual size observed with unaided eye. The total magnification of microscope is calculated by multiplying the magnifying power of the objective lens by that of eye piece .

TERMS AND DEFINITIONS:

Resolving power It is the ability to differentiate two close points as separate. The resolving power of human eye is 0.25 mm The light microscope can separate dots that are 0.25µm apart. The electron microscope can separate dots that are 0.5nm apart. Wavelength of light used is major factor in resolution Shorter wavelength  Greater resolution TERMS AND DEFINITIONS

TERMS AND DEFINITIONS:

Working distance It is the distance between the objective and the objective slide. The working distance decreases with increasing magnification . TERMS AND DEFINITIONS

TERMS AND DEFINITIONS:

TERMS AND DEFINITIONS Numerical aperture(NA) of a lens is the ratio of the diameter of the lens to its focal length. It is an index of the resolving power. NA can be decreased by decreasing the amount of light that passes through a lens. Diameter of the lens

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Eyepiece Body Tube Revolving Nosepiece Arm Objective Lens Stage Stage Clips Coarse Focus Fine Focus Base Diaphragm Light Parts of Microscope MECHANICAL OPTICAL

Base:

Base The bottom of the microscope, used for support Hold this part with one hand when carrying a microscope ← Parts of Microscope ← Supports the tube and connects it to the base The part you hold when you carry the microscope → The long tube that holds the eyepiece and connects the objective

Nosepiece:

Nosepiece Rotating part of the microscope at the bottom of the body tube. It holds the objective lenses ← Nose piece

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The flat platform where you place your slides.  Mechanical stage ← Stage Stage clips : Shiny clips on the top of the stage Holds a slide in place

Knobs:

Large, round knob on the side of the microscope. Either moves the stage or the top part of the microscope up and down → Knobs Small, round knob on the side of the microscope.Used to fine tune the focus after using the coarse adjustment knob →

Condenser & Diaphragm:

Condenser & Diaphragm Produces a cone of light and focuses it on the specimen Regulates the amount of light passing through the slide

):

) Projects or reflects the light so the specimen is easier to see ← Mirror

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Ocular lens also called eyepiece; magnifies an object 10x Objective lenses Objectives Low (4x) Medium (10x) High (40x)

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- Highest magnification - Oil prevents refraction of light outwards and allows it to pass straight in to objective GLASS OIL A B C D E G F FBEG - OIL ABCD - AIR Oil Immersion Objective

Eyepiece:

Eyepiece The lens at the top that you look through. 10X power Forms magnified virtual & erect image → Eye piece

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TYPES (a) Monocular (b) Binocular (c) Trinocular or (a) Huygenian (b) Ramsden (c) Compensating Eye piece

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Always store covered Make sure it’s on a flat surface Always carry with 2 hands. One on the arm and the other on the base Clean only with a soft cloth/tissue Only use lens paper for cleaning Do not force knobs Guidelines for Use Guidelines for use

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Optical system to enhance the contrast of unstained bodies . Specimen appears gleaming bright against dark background Uses Useful in demonstrating Treponema pallidum Leptospira -Campylobacter jejuni-Endospore Dark Field Microscope

Dark field microscope:

Dark field microscope Paramecium Treponema vincenti Volvox and Spirogyra

Phase-contrast microscope :

Phase-contrast microscope First described in 1934 by Dutch physicist Frits Zernike Principle Unstained bacteria have constituents of different refractive index Enhances the contrast between intracellular structures having slight differences in refractive index Advantage -Living cells can be examined in their natural state Phase Contrast Microscope

Phase Contrast Microscope:

Phase Contrast Microscope Special condenser and objective lens which splits the light beam and enhances contrast

Phase contrast microscope:

Phase contrast microscope Phase contrast enables visualization of internal cellular components. Diagnosis of tumor cells . Examination of growth, dynamics, and behavior of a wide variety of living cells in cell culture Uses of Phase Contrast Microscope

Fluorescence Microscopy :

Fluorescence Microscopy Specimen is exposed to ultraviolet, violet, or blue light Stained with fluorochromes Shows a bright image of the object resulting from the fluorescent light emitted by the specimen Fluorescence Microscope

Fluorescence microscope:

Fluorescence microscope Oral cavity Bacillus subtilis

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Some natural sub. exhibit double refraction i.e. they split a beam of light into two, each with its own refractive index value A Nicol prism – polarizer, below condenser lens, other above objective lens, analyzer . Analyzer is rotated until its axis is perpendicular to that of polarizer, no light can pass through the ocular lens resulting in a dark field effect Polarising Microscope

Electron microscope :

Electron microscope The electron microscope uses a beam of electrons rather than visible light. The magnified image is visible on a fluorescent screen and can be recorded on a photographic film. Drawbacks Specimen are killed in order to view the cells or organisms. Images produced by electrons lack color , Are shades of black, gray, and white Electron Microscope

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Transmission Electron Microscope (TEM) Scanning Electron Microscope (SEM) Types of Electron Microscope

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TEM are method of choice for viewing detailed str of cells & viruses. TEM produces image by transmitting electrons through the specimen. Extremely thin slices (20–100 nm) are taken & stained with metals Transmission Electron Microscope

Transmission Electron Microscope:

Projector Lens forms image on Fluorescent viewing screen 2D Image Magnification 10,000 X to 100,000 X Transmission Electron Microscope

Scanning Electron Microscope (SEM) :

Scanning Electron Microscope (SEM) SEM has resolution 1000 times better than Light microscope Can magnify up to 100,000x Uses electrons reflected from the surface of a specimen to create image Produces a 3-dimensional image of specimen’s surface features Scanning Electron Microscope (SEM)

Scanning Electron Microscope (SEM):

Scanning Electron Microscope (SEM) Specimen is placed in vacuum chamber& covered with a thin coat of gold Electron beam scans across specimen & knocks loose showers of electrons that are captured by a detector. An image builds line by line, as in a television receiver.

NEWER MICROSCOPE :

NEWER MICROSCOPE SCANNING PROBE MICROSCOPE Class of Microscope that measures surface features by moving a sharp probe over object surface. Used to visualize atoms and molecules Scanning Tunneling Microscope (STM) Atomic Force Microscope (AFM) Newer Microscope

CONFOCAL SCANNING LASER MICROSCOPE:

CONFOCAL SCANNING LASER MICROSCOPE Uses a laser beam to illuminate a specimen whose image is then digitally enhanced for viewing on a computer monitor. Uses Observing cellular morphology in multilayered specimen Eg . used in diagnosing Ca cervix Evaluation and diagnosis of basal cell carcinoma of skin Confocal Scanning Laser Microscope

Light Vs Electron microscope:

Light Vs Electron microscope

Electron microscope(Facts):

Much more expensive Need more space & require additional rooms for preparation of specimens & processing of photographs The actual image made by electron beam is not visible, would burn eyes if tried to viewed directly Electron beam will eventually burn through the specimen so photos are taken immediately either (micrographs) or on video screen Electron microscope(Facts)