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Cell Fractionation:

Ultracentrifugation:

Process where cells are broken up and the different organelles they contain are separated out
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Before this begins, the tissue is placed in a cold, buffered
solution of the same water potential as the tissue
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1)

tube of filtrate is placed in the centrifuge and spun at slow speed
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3)

The fluid at the top of the tube (supernatant) is removed, leaving
just the sediment of nuclei
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could alter the structure of the organelles or affect the
functioning of enzymes
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Two stages of Cell fractionation:

6)

Process is continued in this also that at each increase of speed, the
next heaviest organelle is sedimented and separated out
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Same water potential as the tissue = to prevent
organelles bursting or shrinking as a result of osmotic
gain or loss of water
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This releases the organelle from the cell
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Magnification:
The magnification of an object is how many times
bigger the image is when compared to the object
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Parts of this
specimen will absorb electrons and make
it appear dark
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All limitations of TEM apply to SEM
but specimens don’t need to extremely thin because electrons
don’t penetrate

Resolving power is 0
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Electrons are scattered on the specimen and the pattern depends on
the contours of the specimen surface

Microscopes

Size of real object = Size of image

Magnification

Resolution / Resolving Power:

The minimum distance apart that two objects can be in order for them to appear
as separate items
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Main limitations: living specimens cannot
be observed because the system works in
Greater resolution means greater clarity
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and even the image isn't in colour
Light microscopes:
Poor resolution as a result of the relatively long wavelength of light
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electron beam has a very short wavelength and the microscope
can therefore resolve objects well = high resolving power

2D images

2
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A scale is etched on the disc
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To calibrate it you need to use a stage micrometer
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Basic SEM has a lower resolving
power



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