Light Microscope

Light microscope

Mechanical & Optical parts

Mechanical parts incl

Base

Stage

Specimen stage

Revolver

Tube & tube holders

Macrometer (Coarse focus)

Micrometer (Fine focus)

Specimen Stage

Metallic plate serving as holder of the slide

What is the opening in the center of the stage for?

Light Transmission

Tube Holders Position (include eyepieces)

Either straight/slanted or monocular/binocular

Slanted at 45-60 degrees

Reflecting Prism function

Changes direction of light

Modern microscopes

Are binocular with 2 eyepieces

Tube length in Modern microscopes?

160-170mm long black cylinder

Modern microscope eyepiece distance

55 - 75mm

Macrometer

Sets distance between specimen and objective lens

Micrometer

Used for fine focus setting

Optical Parts

Serve for illumination and magnification of the specimen

Illumination System

Below the specimen 

Consists of mirror, light source & condenser

Magnification System

Above the specimen

Consists of eyepieces (Objective & Ocular lens)

Light Source

Natural (sunshine) or Artificial (lamp)

Do modern microscopes rely on natural light?

No, as they possess a built-in light source

Condenser

Compound lens system with short focus distance, collects/focuses light on studied object

Objectives (Lens)

Compound lens system with a short focus distance, producing reversed enlarged true image of the object

Consist of several lenses. Most important is plan-convex front lens

Revolving system

Enables viewer to change the objective

Magnification of eyepieces

Between x5 - x25

Total Magnification

Product of multiplication of magnification of individual lenses

Objective lens x Eyepiece

Resolving Power

Minimum distance between 2 points

What improves/increases the resolution?

Use of immersion oil and blue/UV light

What is the maximum magnification and why?

Between x600 - x1000

Above this value image becomes more blurred

Stereo microscope

Include 2 complete light microscopes

Suitable for dissection of smaller anatomical parts (papillae of tongue, microsurgery (eye operations) or pathological diagnostic examinations).

Why are stereo microscopes suitable for microsurgery or dissection of small anatomical parts?

Eyepiece magnification is higher than that of the objective lens

Angle of optical axes of stereo microscope

14 degrees

Illumination of stereo microscope

Either reflective or transmitted or both

Dark Field Microscope Characteristics

Has special condenser that produces dark background

Studied cells appear as illuminated objects against black background

Used to identify bacteria

Condenser of dark field microscope

Blocks & directs light rays

Only allows oblique/peripheral light to reach specimen

Polarization Microscope

Light microscope with 2 polaroid filters:

Polarizator & Analizator

Polarizator

Built in the condenser

Provides polarized light, containing photons

Analizator

Between the objective and ocular lenses (eyepieces)

Either in parallel/perpendicular position to the polarizator

If Analizator is in Parallel position to polarizator?

Light passes through 

Field is bright

If Analizator is in perpendicular position to the Polarizator?

No light passes through

Field is dark

Effect of Anisotropic Materials

Changes vibration plane of polarized light

Causing structures to appear bright against dark background

Fluorescence Microscope Characteristics

Light source is powerful UV lamp

Light passes through 2 filters: one filters light before reaches specimen and other to filter light obtained by specimen

Only fluorescent light emitted by sample is used to form the image

Confocal Laser-scanning microscope Characteristics

Light source: Focused laser beam, moves rapidly in sample

Observe image in single plane of focus, creating sharper image

What is confocal laser-scanning microscope used for?

Studying thicker samples

Electron Microscope Characteristics

Use 5 electromagnetic lenses in its imaging system

Magnification over x1 million at resolving power of 0.1nm

Transmission Electron Microscope (TEM) Components

Vacuum system

Electron optical column

Necessary electronics (lens supplies for focus/deflecting beam & high voltage generator

TEM Characteristics

Light source: Electron gun, built into column

Electromagnetic lenses instead of glass lenses

Image appears on fluorescent screen

TEM emitted electrons

Entire electron path has to be under a vacuum

Why does the entire electron path have to be under a vacuum?

So image can be viewed through a window in projection chamber

Electron gun components

Filament

Anode

Wehnelt cylinder

These 3 form triode which is a very stable source of electrons

Tungsten Filament

Hairpin shaped

Heated to ¬2700c

Wehnelt Cylinder

Bunches electrons into finely focused point

Electromagnetic lenses comprises?

Condenser System

Objective lens

Imaging lenses

Condenser lens system

Focus electron beam onto specimen under investigation

Objective lens

Produces an image of the specimen

Imaging lens

Magnifies the image produced by objective lens and projects it onto the fluorescent screen

From filament - fluorescent screen what happens to the beam?

It passes through a series of apertures with different diameters

Apertures

Stop electrons which do not contribute to the process of image formation