APPLICATIONS OF THE MICROSCOPE

APPLICATIONS OF THE MICROSCOPE

APPLICATIONS OF THE MICROSCOPE

• Brightfield Microscopy

• Darkfield Microscopy

• Phase Contrast Microscopy

• Electron Microscopy

  - Transmission Electron Microscope (TEM)

  - Scanning Electron Microscope (SEM)

BRIGHTFIELD MICROSCOPY

- The simplest of all the optical microscopy illumination techniques

BRIGHTFIELD MICROSCOPY

- Specimen’s image appears dark against a bright background

BRIGHTFIELD MICROSCOPY

- Light source is usually tungsten lamp

BRIGHTFIELD MICROSCOPY

Generally used with compound microscopes

BRIGHTFIELD MICROSCOPY

Terms:
1. RESOLVING POWER
2. WHITE LIGHT
3. NUMERICAL APERTURE
4. TOTAL MAGNIFICATION

RESOLVING POWER

- Ability to distinguish between two adjacent points
- Depends on two factors: White light & Numerical aperture

WHITE LIGHT

- Composed of a mixture of colored lights of various wavelengths

NUMERICAL APERTURE

- Expression relating to the cone of light that is delivered to the specimen by the condenser and gathered by the objective

TOTAL MAGNIFICATION

• the ocular magnification multiplied by the objective magnification

PHASE CONTRAST MICROSCOPY

- Uses a special optic system which converts differences in phases into differences in intensity such that some structures appear darker than the others

PHASE CONTRAST MICROSCOPY

- Reveals the gross details of the internal structures in a living cell

DARKFIELD MICROSCOPY

Objects appear brilliantly illuminated against a black background

DARKFIELD MICROSCOPY

Used to demonstrate spirochetes which are difficult to observe in transmitted light

2 TYPES OF ELECTRON MICROSCOPY

1. TEM (TRANSMISSION ELECTRON MICROSCOPE)

2. SEM (SCANNING ELECTRON MICROSCOPE)

TEM (TRANSMISSION ELECTRON MICROSCOPE)

• Develops an image resulting from variable electron density of the specimen interposed in the electron beam

TEM (TRANSMISSION ELECTRON MICROSCOPE)

• Produces monotone, two-dimensional, high magnified images

TEM (TRANSMISSION ELECTRON MICROSCOPE)

• Resolving power is 0.001 um, which is 200X that of light microscopes

TEM (TRANSMISSION ELECTRON MICROSCOPE)

• Specimens must be fixed, stained, and dried

TEM (TRANSMISSION ELECTRON MICROSCOPE)

• Used to observe internal ultra-structural detail of cells and observations of viruses or small bacteria

SEM (SCANNING ELECTRON MICROSCOPE)

• Has a practical limit of resolution of 0.005um or five-fold less than that of TEM

SEM (SCANNING ELECTRON MICROSCOPE)

Produces monotone, tri-dimensional image by detection at a 90 degrees angle of secondary electrons emitted from the specimen surface as a result of bombardment by the primary electron beam

SEM (SCANNING ELECTRON MICROSCOPE)

• Used to observe the surface details of structures

MICROSCOPE HANDLING AND STORAGE

1. When work is completed, lower the stage, remove the slide, rotate in the lowest power objective, wrap the cord loosely around the base and cover with a dust cover. Take care not to wrap the cord around a hot (built in) light source.

2. Always keep your microscope covered when not in use. Optics and mechanical parts must be protected from dust.

3. Always move the microscope with one hand under the base and the other hand gripping the arm or frame.

4. Keep microscopes away from vibration, moisture, high temperatures and direct sunlight.

5. Never store microscopes in chemical storage areas as corrosive fumes may damage metal and lenses.

MICROSCOPE MAINTENANCE

1. Treat lenses with great care as they can be easily scratched. Never use anything abrasive.

2. When cleaning lenses, first blow away any dust with a blower brush then use lens tissue and lens cleaning fluid to clean the objectives and eyepieces. Do not use paper towel or regular tissues, as they will scratch the lens. Do not use other solvents.

3. Do not remove eyepieces or objectives from their location but clean only their external surfaces.

4. Remove immersion oil from the 100x objective immediately after use with lens tissue and lens cleaning fluid.

5. Wipe dust off the body of the microscope with a damp cloth