Lab 2- Introduction to Microscopes

Bright field Microscope

1. Requires staining

2. Requires heat fixation to secure object on slide during oil immersion

3. Staining and heat fixing can damage cell morphology and can kill the cells

Compound Light Microscope

1. Light is passed through the slide

2. Light is not scattered by the object and enters the lens

3. Objective lends

4. Ocular lens

Objective lens

Produces real image of object inside microscope (Objective magnification)

Ocular lens

Produces virtual image seen by observer. (Objective x ocular magnification)

Dark Field Microscopy

1. Light is passed through the slide from a side angle

2. Light scattered by object enters the lens

3. Light not scatted by the object is not captured

4. Advantage: Can be used for observing live and unstained objects

5. Disadvantage: Attachments increase microscope cost

Dark Field Microscope

Advantage: Can be used for observing live and unstained objects

Phase-Contrast Microscope

1. Uses alternate phasing of light wavelengths to hit object at different angles

2. Used for overserving live and unstained objects

Phase-Contrast Microscope

Advantage: Phase shift of light show subtle differences in internal density, showing more features and details

Fluorescence Microscope

1. Uses photoluminescence to observe objects

2. Monochromatic (single wavelength) light is used to excite fluorochromes on object

3. The sample is often treated with antibodies with fluorochromes that bind to cell components

4. The monochromatic light excites the fluorochromes, causing them to emit photons of light of a different wavelength

Fluorescence Microscope

Advantage: Specific features of the object can be identifies and light does not have to pass through specimen

Electron Microscope

1. Electrons are fires at the sample from a cathode

2. Image is magnified through a magnetic coil and captured on a photographic plate

3. Transmission or scanning techniques can show interior or surface, respectively

Electron Microscope

Advantage: High magnification (10^6 x) and resolution

Electron Microscope

Disadvantage: Cost and time, Sample is destroyed in process

Interpupillary adjustment

Adjust distance between ocular lenses depending on distance between your eyes

Diopter adjustment ring

Allows left eye to be focused independently of right eye

Ocular lens

Eye piece lens; magnifies 10x

Objective lens of 4x

Scanning

Objective lens of 10x

Low Power

Objective lens of 40x

High Power

Objective lens of 100x

Oil Immersion

Total Magnification

Ocular magnification * Objective Magnification

Stage

Place for slide/specimen to lay

Slide Holder/Stage Clip

Holds slide and specimen for the mechanical stage

Lamp or Illuminator

Illuminates specimen

Base

Supports microscope and used to carry microscope

Stage Adjustment (X-Y Axis)

Moves stage right or left and backwards or forwards

Iris Diaphragm

Regulates amount of light that reaches specimen

Magnification

Increasing the virtual size of the specimen

Resolution

The ability of a lens to distinguish between 2 adjacent points

Resolving Power (R.P)

The minimum distance 2 points can be apart, and the lens still has resolution

Contrast

Difference between lights and dark (Low contrast=dark)

Refraction

Bending of light as it moves from one medium to an other

Numerical aperture (N.A)

Light capturing ability of lens

Working distance

Distance between objective lens and specimen

Depth of field

Thickness of specimen that is in focus at one time (At higher magnification you can only focus on one thread below)

Range of field

The area of the slide shown in the observed image

Increases

Working distance, depth of field, and range of field all decrease as magnification ____

Parfocal

A microscope that stays in focus when the magnification (lens) change