01 Microscope in cell studies

1. CELL STRUCTURECELL STRUCTURE
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2. Early daysEarly days
Hans and Zacharias Janssen of Holland in the Hans and Zacharias Janssen of Holland in the
1590’s created the “first” compound 1590’s created the “first” compound
microscopemicroscope
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3. 1665 - Robert Hooke 1665 - Robert Hooke
discovered and described the discovered and described the
fundamental unit of all living fundamental unit of all living
things (cells) by examining things (cells) by examining
thin slices of corkthin slices of cork
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4. 1674 - The first man to 1674 - The first man to
witness a live cell witness a live cell
under a microscope under a microscope
was Anton Van was Anton Van
Leeuwenhoek, Leeuwenhoek,
describing the algae describing the algae
SpirogyraSpirogyra and named and named
the moving organisms the moving organisms
animalcules, meaning animalcules, meaning
"little animals" "little animals"
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6. 1838 – Matthias Schleiden suggested that all 1838 – Matthias Schleiden suggested that all
plants are made of cellsplants are made of cells
1839 – Theodor Schwann suggested the same 1839 – Theodor Schwann suggested the same
for animalsfor animals
General General cell theorycell theory emerged: emerged:
““the basic unit structure and function of all living the basic unit structure and function of all living
organisms is the cell”organisms is the cell”
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7. 1855 – Rudolph Virchow’s theory: “all cells arise 1855 – Rudolph Virchow’s theory: “all cells arise
from pre-existing cells by cell division”from pre-existing cells by cell division”
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8. Unit of Measurements in Cell Unit of Measurements in Cell
StudiesStudies
metremetre mm= 1 m= 1 m
millimetremillimetremmmm= 10= 10
-3-3
m m
micrometremicrometreµmµm= 10= 10
-6-6
m m
nanometrenanometre nmnm= 10= 10
-9-9
m m
picometrepicometre pmpm= 10= 10
-12-12
m m
angstromangstrom AA= 10= 10
-10-10
m (obsolete) m (obsolete)
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10. Magnification and ResolutionMagnification and Resolution
Magnification Magnification
- the number of times larger an image is - the number of times larger an image is
compared with the real size of the objectcompared with the real size of the object
- magnification = size of image- magnification = size of image
actual size of specimenactual size of specimen
ResolutionResolution
- the ability to distinguish between two separate - the ability to distinguish between two separate
points points
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11. The Microscope in Cell StudiesThe Microscope in Cell Studies
2 different types of microscopes:2 different types of microscopes:
i)i)light microscopelight microscope – uses light as a – uses light as a
source of radiationsource of radiation
ii)ii)electron microscopeelectron microscope – uses – uses
electrons electrons
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12. Light microscopeLight microscope
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13. Carry a Microscope CorrectlyCarry a Microscope Correctly

14. Images Produced by Light Images Produced by Light
MicroscopesMicroscopes
Amoeba Streptococcus bacteria Anthrax bacteria
Human cheek cells
Plant cells
Yeast cells

15. Electron microscopeElectron microscope
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16. Images Produced by Electron Images Produced by Electron
MicroscopesMicroscopes
Cyanobacteria
(TEM) Lactobacillus
(SEM)
Campylobacter
(SEM)
Deinococcus
(SEM)
House ant
Avian influenza
virus
Human eyelash
Yeast

17. Electrons – negatively charged particles Electrons – negatively charged particles
which orbit the nucleus of an atomwhich orbit the nucleus of an atom
When electrons gain high energy, they When electrons gain high energy, they
escape from their orbits. Free electrons escape from their orbits. Free electrons
behave like electromagnetic radiationbehave like electromagnetic radiation
Short wavelength (greater energy, shorter Short wavelength (greater energy, shorter
wavelength)wavelength)
Advantage of using electrons for Advantage of using electrons for
microscopy:microscopy:
Wavelength extremely shortWavelength extremely short
Negatively charged (can be focused easily Negatively charged (can be focused easily
using electromagnets)using electromagnets)
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18. Electromagnetic spectrum – whole range of wavelengthsElectromagnetic spectrum – whole range of wavelengths
The longer the electromagnetic waves, the lower their The longer the electromagnetic waves, the lower their
frequencyfrequency
The greater the energy, the shorter the wavelengthThe greater the energy, the shorter the wavelength
The limit of resolution is about ½ the wavelength The limit of resolution is about ½ the wavelength
of the radiation used to view the specimen of the radiation used to view the specimen (if an (if an
object is any smaller than half the wavelength of the object is any smaller than half the wavelength of the
radiation used to view it, it cannot be seen separately radiation used to view it, it cannot be seen separately
from nearby objects)from nearby objects)
Best resolution using microscope that uses visible light is Best resolution using microscope that uses visible light is
200 nm, since shortest wavelength of visible light is 400 200 nm, since shortest wavelength of visible light is 400
nm (violet light). Smaller objects can never be seen nm (violet light). Smaller objects can never be seen
using light (ribosomes – 22 nm)using light (ribosomes – 22 nm)
If an object is transparent it will allow light waves to If an object is transparent it will allow light waves to
pass through it and will still not be visiblepass through it and will still not be visible
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The electromagnetic spectrumThe electromagnetic spectrum

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21. Beam of electrons passed Beam of electrons passed throughthrough the the
specimen before being viewedspecimen before being viewed
Only electrons that are Only electrons that are transmittedtransmitted
(pass through the specimen) are seen(pass through the specimen) are seen
Allows to see thin sections of specimens Allows to see thin sections of specimens
(inside the cell)(inside the cell)
Transmission Electron MicroscopeTransmission Electron Microscope
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23. Electron beam is used to scan the Electron beam is used to scan the surfacessurfaces of of
structures and only the structures and only the reflectedreflected beam is beam is
observedobserved
Surface structures can be seenSurface structures can be seen
Great depth of field is obtained so that much of Great depth of field is obtained so that much of
the specimen is in focus at the same timethe specimen is in focus at the same time
Cannot achieve same resolution as a Cannot achieve same resolution as a
transmission electron microscopetransmission electron microscope
Scanning Electron MicroscopeScanning Electron Microscope
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25. Viewing sample with the electron Viewing sample with the electron
microcopemicrocope
Electron beam projected onto a fluorescent Electron beam projected onto a fluorescent
screen (areas hit by electrons shine brightly screen (areas hit by electrons shine brightly
giving overall a ‘black and white’ picture)giving overall a ‘black and white’ picture)
Stains used to improve contrast contain heavy Stains used to improve contrast contain heavy
metal atoms which stop the passage of electronsmetal atoms which stop the passage of electrons
Electron beam must be in vacuum to avoid Electron beam must be in vacuum to avoid
collision with air moleculescollision with air molecules
Specimen must be dehydrated because water Specimen must be dehydrated because water
boils at room temperature in vacuum (only dead boils at room temperature in vacuum (only dead
material can be examined)material can be examined)
Great efforts to preserve material in life-like Great efforts to preserve material in life-like
state when preparing specimenstate when preparing specimen
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26. Comparison of advantages and disadvantages of the light and electron Comparison of advantages and disadvantages of the light and electron
microscopemicroscope
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LIGHT MICROSCOPELIGHT MICROSCOPE ELECTRON MICROSCOPEELECTRON MICROSCOPE
AdvantagesAdvantages DisadvantagesDisadvantages
Cheap to purchaseCheap to purchase Expensive to purchaseExpensive to purchase
Cheap to operate – uses a little electricity where Cheap to operate – uses a little electricity where
there is a built-in light sourcethere is a built-in light source
Expensive to operate – requires up to 100 000 Expensive to operate – requires up to 100 000
volts to produce the electric beamvolts to produce the electric beam
Small and portable Small and portable Very large and must be operated in special roomsVery large and must be operated in special rooms
Unaffected by magnetic fieldsUnaffected by magnetic fields Affected by magnetic fieldsAffected by magnetic fields
Preparation of material is relatively quick and Preparation of material is relatively quick and
simple, requiring only a little expertisesimple, requiring only a little expertise
Preparation of material is lengthy and requires Preparation of material is lengthy and requires
considerable expertise and sometimes complex considerable expertise and sometimes complex
equipmentequipment
Material rarely distorted by preparationMaterial rarely distorted by preparation Preparation of material may distort itPreparation of material may distort it
Living as well as dead material may be viewedLiving as well as dead material may be viewedA high vacuum is required and living material A high vacuum is required and living material
cannot be observedcannot be observed
Natural colour of material can be observedNatural colour of material can be observed All images are in black and whiteAll images are in black and white
DisadvantagesDisadvantages AdvantagesAdvantages
Magnifies objects up to 1500XMagnifies objects up to 1500X Magnifies objects over 500 000XMagnifies objects over 500 000X
Can resolve objects up to 200 nm apartCan resolve objects up to 200 nm apart Has a resolving power for biological specimens of Has a resolving power for biological specimens of
around 1 nmaround 1 nm
The depth of field is restrictedThe depth of field is restricted It is possible to investigate a greater depth of It is possible to investigate a greater depth of
fieldfield

27. GraticuleGraticule
Stage micrometerStage micrometer
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