MICROSCOPES AND RESEARCH

TOOLS IN CELL BIOLOGY

 Lecture Outline

• Types of microscopes and their importance

• Types of other experimental tools
 What are the
types of
microscopy
available and their
functions?
 The Emergence of Modern Cell Biology

• Three historical strands weave together into

modern cell biology, each with important
contributions to understanding cells

• The cytology strand focuses mainly on

cellular structure and emphasizes optical
techniques

• The biochemistry strand focuses on cellular

function

• The genetics strand focuses on information

flow and heredity
Why Microscopy Field is Important?
 Intestinal cell structure

Consistent array
of proteins,
including an
electrically
powered ATP-
Microvilli  synthesizing
Long machine (ball on
projections a stick structure)
that facilitate
nutrients
absorption
Defined pattern
of internal
membranes
Actin (protein)
 Filamentous
skeleton
allowing the
outward Mitochondria 
projection Provide energy
from apical cell for cellular
surface processes
 Microscope Types

• Types of microscopes
1. Bright-field light
microscopy
2. Phase contrast light
microscopy
3. Fluorescence microscopy
4. Polarizing microscopy
5. Electron microscopy
 Light Microscopy

• The light microscope was the

earliest tool of cytologists
• Allowed identification of
organelles within cells
• Organelles are membrane-
bound structures, such as
nuclei, mitochondria and
chloroplasts
 Bright-Field Light Microscopy

• Bright-field  Classical microscopes in which

the sample is illuminated from the back and the
image is formed due to the absorbing
properties of the imaged objects
• The technique is best suited for utilization with
fixed, stained specimens or other kinds of
samples that naturally absorb significant
amounts of visible light
• Images produced with bright-field illumination
appear dark and/or highly colored against a
bright, often light gray or white background
 Bright-Field Light Microscopy

• For BF light microscopy, some sample preparation steps

(fixing, staining, embedding in plastic) may distort tissues 
Microscopy techniques to view live cells
 Visualizing Live Cells

1. Phase contrast/differential
interference contrast microscopy

2. Fluorescence microscopy

3. Confocal scanning
 Phase Contrast Light Microscopy

• Optical microscopy technique that enhances contrasts of

transparent and colorless objects by influencing the optical
path of light  Specimen appearing as dark object
 Fluorescence Microscopy

• An optical microscope that uses fluorescence

and phosphorescence instead of, or in
addition to, reflection and absorption to study
properties of organic or inorganic substances
• Demonstrate naturally occurring fluorescence
material and non-fluorescence
materials/microbes with fluorescent dye
staining  To show locations of substances in
the cell
• UV light used for illumination
• Confocal microscopy  Special type of
fluorescence microscopy that uses a laser
beam to illuminate a single plane of a
fluorescently labeled specimen
 Polarizing Microscopy

• Demonstrate birefringence
• Birefringence objects rotate light
rays  Appearing bright in dark
background
• Eg. Amyloid, foreign body

Polarized microscopy of Congo red–

stained myocardium showing yellow-
green birefringence of amyloid deposits
 Electron Microscopy

• The electron microscope, using a beam of

electrons rather than light, was a major
breakthrough for cell biology
• Used to study ultrastructural details of
tissues and cells
• Tissues fixed in 4% glutaraldehyde at 4oC
for 4 hours  Ultra-thin microsections
with 100nm thickness cut using diamond
knives
• Limit of resolution  Around 0.1-0.2 nm
• High resolving power  50000x to
100000X
 Electron Microscopy
• Transmission electron microscopy (TEM)
 Electrons transmitted through specimen
 Visualizes cell cytoplasm and organelle
 Ultra-thin sections required
 Interprets atomic properties
 Gives 2D image

• Scanning electron microscopy (SEM)

 Study of cell surface  Electrons deflected
from outer surface
 Can be used for fluorescent antibody
techniques
 Gives 3D image
 Electron Microscopy

These images
are from TEM
or SEM?
What are the
other
experimental
tools to study
cell?
 Other Research Tools in Cell Biology

• Research in laboratories may be

1. In vitro  Using live cells or purified chemicals
and cellular components
2. In vivo  Using live organisms
3. In silico  Using computer analysis of large
amounts of data
 In vitro Research Tools

PC3 (prostate) cell line culture Cell fractionation

 In vivo Research Tools

SCID mice

White mice (Balb/C; ICR)

Zebra fish
C57BL/6 mice

Rabbit
In silico Research Tools