1. Concept: Biologists use microscopes and the tools of biochemistry to study cells
first compound microscope – Zacharias Jansen in 1590
three important parameters of microscopy
Magnification: the ratio of an object’s image size to its real size
Resolution: the measure of the clarity of the image, or the minimum distance of two distinguishable points
inversely related to the wavelength of the radiation a microscope uses
Contrast: visible differences in parts of the sample

2. Studying Cells: Microscopy
1 cm
1 mm
100 m
10 m
1 m
100 nm
10 nm
1 nm
0.1 nm
Atoms
Small molecules
Lipids
Proteins
Ribosomes
Viruses
Smallest bacteria
Mitochondrion
Most bacteria
Nucleus
Most plant and animal cells
Human egg
Frog egg
Chicken egg
Length of some nerve and muscle cells
Human height
Unaided eye
Light microscopy
Electron microscopy
Super- resolution microscopy
in a light microscope (LM) - visible light is passed through a specimen and then through glass lenses
the lenses refract (bend) the light - so that the image is magnified
in a simple microscope - there is one lens for magnification
e.g. magnifying lens
in compound microscopes – more than one lens
ocular and objective lenses
improves resolution and allows for more than one magnification
LMs can magnify effectively to about 1,000 times the size of the actual specimen
this allows for individual cells within a tissue to be visualized

3. Brightfield
(unstained specimen)
(stained specimen)
50 m
Differential-interference- contrast (Nomarski)
Fluorescence
10 m
Light Microscopy (LM)
Phase-contrast
various techniques enhance contrast of a LM and enable cell components to be stained or labeled
BUT - most subcellular structures, including organelles, are too small to be resolved by an LM
LMs cannot resolve detail finer than 0.2um - regardless of magnification
Figure 6.3 Exploring: Microscopy

4. to improve resolution and magnification – allowing for imaging of subcellular structure - development of two other microscopes in the 1950s
called electron microscopes (EMs)- use a focused beam of electrons rather than light
resolution increase – due to the shorter wavelength of the electron beam
two types:
1. Transmission Electron Microscope (TEM)
2. Scanning Electron Microscope (SEM)

5. Blood cells
1. Scanning electron microscopes (SEMs) – electron beam is focused onto the surface of a subject
providing images that look 3D
SEM electron beam excites the electrons of the gold on the subject’s surface
several kinds of electrons are produced
these electrons are detected by the scope and projected onto a video screen as a magnified image that appears 3D
can be colorized
Pollen grains

6. 2. Transmission electron microscopes (TEMs) focus a beam of electrons through a specimen
subject is sliced into a very thin layer
so TEMs are used mainly to study the internal structure of cells
subject is stained with heavy metals that adhere to the internal structures of the cell
so some parts of the cell become more electron dense than others
the electron beam passes through those less dense and scattered/reflected by the more dense regions
the electrons that pass through hit a piece of film negative or hit a detector for displaying the image