Methods Of Studing Cells (Microscopes) Flashcards
Define magnification
Tells you how many times bigger the image produced by the microscope is that the real life object
Define resolution
The ability to distinguish between two objects that are close together
How to convert millimetres to micrometres (mm - um)
x1000
How to convert micro meters into nanometres
x1000
Magnification equation
Magnification = Size of image / Size of real object
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A M
What optical microscopes use to form an image
Light
What is the maximum resolution of a light microscope
0.2 micrometers (um)
What is the maximum magnification of an optical microscope
x 1500
What organelles can’t be seen with an optical microscope
Ribosomes, endoplasmic reticulum, lysosomes
What do electron microscope is used for an image
Electrons
Which has higher resolution TEM or SEM
TEM
Which has greater magnification electron or light microscope
Electron
What does the images that electron microscopes produce look like
Black and white
(Can be coloured by computer) 
What are the two types of electron microscope
Transmission electron microscope’s (TEMs)
Scanning electron microscope’s (SEMs)
How does a transmission electron microscope work
Use electromagnets to focus a beam of electrons which is transmitted through the specimen
Advantages of TEMs
High resolution
Can see internal structures
Disadvantages of TEMs
Thin specimens only
Can’t view living organisms - uses vacuum so no water
No coloured image
What is an artefact in microscopy
Looks like a real structure but is the result of preserving or staining a specimen
What does the image from TEM look like and WHY?
Denser parts absorb more electrons- look darker 
How does a scanning electron microscope work
They scan a beam of electrons across the specimen
The electrons knock off the specimen and collected by cathode to form an image
Advantages of SEM
Can be used as thick specimens
Produce 3-D image
Disadvantages of SEM
Lower resolution than TEM
Cannot be used to observe live specimens
What are the steps of cell fractioning (3)
Homogenisation
Filtration + cold ionic buffer etc
Ultracentrifugation
What does Homogenisation mean
Breaking up of cells
What is done to the sample of the tissue in homogenisation
Placed in an cold,isotonic, buffer solution.
Why is the solution ice-cold
To reduce the activity of enzymes which break down organelles
Why is the solution isotonic
Same water potential as the cell to prevent osmosis/lysis
Why is the solution of buffer
To prevent organelle proteins from becoming denatured
What is done with the solution once prepared ( homogenisation)+ what does this achieve /do
Added to a homogeniser which grinds up the cell
Breaks the plasma membrane releases the organelles into the homogenate ( solution)
What is done during filtration
The homogenate is filtered through a gauze
separate large debris
organelles passed through
What is the pellet
The heavy organelles which settle at the bottom forming a thick sediment
What happens during ultracentrifugation
Put in a tube
Spun in a centrifuge
Supernatant is put in a new tube
spin again
Repeat
What is a supernatant
The remaining organelles which are suspended in the fluid above the sediment
Describe how you’d make a temporary mount of a piece of plant tissue to observe the position of the starch grains in the cell when using an optical microscope
Add drop of water to slide
Add thin section of cell onto slide
Stain with iodine
Lower cover slip using mounted needle
Order of cell separation in cell fractioning
5 layers
1) nuclei
2) mitochondria
3) Lysosomes
4) ERs + Golgi apparatus
5) ribosomes
Why do you
A) push
B)not push sideways
On a microscopic slide
A- spread tissue
B- avoid cells rolling together
Why would an optical microscope not be able to identify an organelle (2 marks)
Low resolution
Light has longer wavelength
Give an example of why you may not be able to see the structure inside a cell
Not stained
How can chloroplasts be isolated from leaves
Homogenisation and filter leaves
Place in cold, isotonic, buffer solution solution
Spin in centrifuges and remove cell debris
Spin at higher speed and remove chloroplasts
Suggest why you cannot see a nucleus in a image
The image is clear and fairly magnified
Not stained
Benefits of TEMs over SEMs
Higher resolution see internal structures
Write out unit conversion from mm to nm
Look on posit for answer
How to use graticule x3
Measure using eyepiece
Calibrate eyepiece gratified against stage ruler
Take number of measurements to calculate mean
