3.1 & 3.2 Methods of studying Cells and The Electron Microscope
Description
AS - Level Biology (3 - Cell Structure) Mind Map on 3.1 & 3.2 Methods of studying Cells and The Electron Microscope, created by Bee Brittain on 31/03/2016.
3.1 & 3.2 Methods of studying
Cells and The Electron Microscope
Optical Microscopes
Use light to form an image
Maximum resolution of about 0.2micrometres
So you can't use an optical microscope to view
organelles smaller than 0.2 micrometres. This
includes ribosomes, ER and lysosomes. You may
see mitochondria, but not in peprfect detail.
Maximum useful magnification of about x1500
Using a slide to view specimens
1) add a small drop of water onto
the slide then use tweezers to place
a thin section of your specimen on
top of the water drop
2) Add a drop of stain to highlight
objects in the cell (e.g Iodene is used
for starch)
3) Finally, add a cover slip. Careful for air bubbles!
Transmission Electron Microscope (TEM)
Use electromagnets to focus a beam on
electrons which is then transmitted
through the specimen
Electron beam has a very short wavelength
High resolving power of 0.1nm
Denser parts of specimen absorb more electrons = darker on image
High resolution images = see internal
structure of organelles such as
chloroplasts
Only can be used on thin specimens
Limitations
Whole system is in a vacumn = specimens must be dead
Specimen has to undergo a complex staining process = can be damaged, not true colour.
Specimen has to be extremely thin
Image may contain artefacts ( things resulting from the way the
specimen is prepared) = foreign objects on end photomicrograph
Scanning Electron Microscope (SEM)
Scan a beam of electrons across the specimen
knocks off electrons from the specimen which
are gathered in a cathode ray tube to form an
image
Images show surface of specimen and so they can be 3D
They have lower resolution than a TEM
20nm
The Limitations of a SEM are very similar to a
TEM, however the specimen doesn't need to be
extremely thin as electrons do not penetrate it
Magnification is Size
Magnification = Image size/Actual size
Magnification is how much bigger the image is than the specimen
Resolution is detail
Resolution is how detailed the image is, and how well a microscope
distinguishes between two points that are close together
Cell Fractionation and Ultracentrifugation
If you want to look at some organelles under an electron microscope,
you'd first need to separate them from the rest of the cell = cell
fractionation.
1) Homogenisation - Breaking up the cells
Cells are broken up by a homogeniser (blender). This breaks up the plasma
membrane and releases the organelles into solution. The solution is called
homogenate.
Solution has to be ice-cold =
reduce activity of enzymes that
break down organelles
Solution has to be Isotonic = same concentration of
chemicals as the cells being broken down/same water
potential as tissue to prevent organelles bursting/shrinking.
A buffer solution should also
be added to homogenate to
maintain pH as a change in pH
could alter the structure of the
organelles
2) Filtration - Getting rid of the big bits
The homogenate is filtered through a gauze to seperate any large
cell debris or tissue debris from the organelles
The organelles are much smaller than the debris so they pass through the gauze
3) Ultracentrifugation - Separating the organelles
Separated in a machine called the centrifuge
The tube of filtrate is placed in the centrifuge and spun at low speed
The heaviest organelles, the nuclei, are forced to the bottom of the tube and form thing sediment
Fluid at the top of the tube (supernatant) is removed, leaving just the sediment of nuclei
The supernatant is transferred to another tube and spun in the centrifuge at a faster speed than before
Next heaviest organelles (e.g mitochondria) forced to the bottom of the tube
The process is continued in this way so with each increase in speed the next heaviest organelle is sedimented and separated