3.1 & 3.2 Methods of studying Cells and The Electron Microscope

Bee Brittain
Mind Map by Bee Brittain, updated more than 1 year ago
Bee Brittain
Created by Bee Brittain about 4 years ago
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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 03/31/2016.

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3.1 & 3.2 Methods of studying Cells and The Electron Microscope
1 Optical Microscopes
1.1 Use light to form an image
1.2 Maximum resolution of about 0.2micrometres
1.2.1 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.
1.3 Maximum useful magnification of about x1500
1.4 Using a slide to view specimens
1.4.1 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
1.4.2 2) Add a drop of stain to highlight objects in the cell (e.g Iodene is used for starch)
1.4.3 3) Finally, add a cover slip. Careful for air bubbles!
2 Transmission Electron Microscope (TEM)
2.1 Use electromagnets to focus a beam on electrons which is then transmitted through the specimen
2.1.1 Electron beam has a very short wavelength
2.1.1.1 High resolving power of 0.1nm
2.2 Denser parts of specimen absorb more electrons = darker on image
2.3 High resolution images = see internal structure of organelles such as chloroplasts
2.4 Only can be used on thin specimens
2.5 Limitations
2.5.1 Whole system is in a vacumn = specimens must be dead
2.5.2 Specimen has to undergo a complex staining process = can be damaged, not true colour.
2.5.3 Specimen has to be extremely thin
2.5.4 Image may contain artefacts ( things resulting from the way the specimen is prepared) = foreign objects on end photomicrograph
3 Scanning Electron Microscope (SEM)
3.1 Scan a beam of electrons across the specimen
3.1.1 knocks off electrons from the specimen which are gathered in a cathode ray tube to form an image
3.2 Images show surface of specimen and so they can be 3D
3.3 They have lower resolution than a TEM
3.3.1 20nm
3.4 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
4 Magnification is Size
4.1 Magnification = Image size/Actual size
4.2 Magnification is how much bigger the image is than the specimen
5 Resolution is detail
5.1 Resolution is how detailed the image is, and how well a microscope distinguishes between two points that are close together
6 Cell Fractionation and Ultracentrifugation
6.1 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.
6.1.1 1) Homogenisation - Breaking up the cells
6.1.1.1 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.
6.1.1.1.1 Solution has to be ice-cold = reduce activity of enzymes that break down organelles
6.1.1.1.2 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.
6.1.1.1.3 A buffer solution should also be added to homogenate to maintain pH as a change in pH could alter the structure of the organelles
6.1.2 2) Filtration - Getting rid of the big bits
6.1.2.1 The homogenate is filtered through a gauze to seperate any large cell debris or tissue debris from the organelles
6.1.2.1.1 The organelles are much smaller than the debris so they pass through the gauze
6.1.3 3) Ultracentrifugation - Separating the organelles
6.1.3.1 Separated in a machine called the centrifuge
6.1.3.2 The tube of filtrate is placed in the centrifuge and spun at low speed
6.1.3.2.1 The heaviest organelles, the nuclei, are forced to the bottom of the tube and form thing sediment
6.1.3.2.1.1 Fluid at the top of the tube (supernatant) is removed, leaving just the sediment of nuclei
6.1.3.2.1.1.1 The supernatant is transferred to another tube and spun in the centrifuge at a faster speed than before
6.1.3.2.1.1.1.1 Next heaviest organelles (e.g mitochondria) forced to the bottom of the tube
6.1.3.2.1.1.1.1.1 The process is continued in this way so with each increase in speed the next heaviest organelle is sedimented and separated
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