Cell Biology – Optical Methods

Textbook: Unit 2

What you need to know!

• The 3 pillars of Cell Theory and their importance.

• Different types of microscopes used by biologists (especially the Electron Microscopes)

Important Discoveries

1. Light microscope 1630• Anton Leeuwenhoek

2. The cell 1665• Robert Hooke

3. Cell Theory 1839• Matthias Schleiden, Theodor Schwann

Cell Theory

1. All living things are made out of one or more cells

• Unicellular: prokaryotes (i.e. bacteria), some eukaryotes

• Multicellular

2. Cells are the basic units of structure and function in living things

a. All body parts are made out of or by cells

b. All functions performed by organisms exist on the cellular level (i.e. breathing, moving, digestion, and reproduction)

c. Organisms have organs, cells have organelles

Cell Theory

3. All cells come from other cellsa. Cell division (asexual)

b. Sperm/egg (sexual)

c. No spontaneous generation

A. Technical terms:

1. Magnifying Power: how much a microscope can magnify

• Depends on the lenses used and the wavelength of the radiation

• Expressed as “x”• Example: 20x (20 times)

2. Resolving Power: the clarity of the image

• The distance between 2 points that can be clearly separated

B. Compound Light Microscope

• Visible light is the radiation source

• Glass lenses are used for magnification

• Images are in color

• Includes: two or more lenses plus a light source

Lenses: Magnification:Eyepiece 10x - 20x

Objective lenses: 4x (low power)

10x (high pwr 1)

40x (high pwr 2)

B. Compound Light Microscope

• Total magnification = lens 1 x lens 2

• Maximum magnification = 1,000x

C. Electron Microscope

• Electron beam is the radiation source

• Lenses are electromagnets

• Specimen is prepared (frozen/dead)

• Images are enhanced by electronics and computers

• Images are black and white

• Electron Microscope (EM) maximum magnification = 500,000x

• 2 types of EM

1. Transmission Electron Microscope (TEM)• Electron beam goes through thin

slices of the specimen

2. Scanning Electron Microscope• Electron beam scans over surfaces of

the entire specimen or broken up specimen (landscape)

• 3D image

Cell Fractioning

1. Homogenize cells

2. Centrifugation the homogenate (spin at 10,000 rpm)

3. Organelles will be in the pellet (bottom of tube)– Heaviest organelles first

Cell Fractioning