Methods of studying cells Flashcards

1
Q

Describe what is meant by cell fractionation

A

The process in which organelles are separated

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2
Q

What are the 2 parts of cell fractionation called

A
  • Homogenisation: cells are broken apart
  • Centrifugation: the organelles are separated
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3
Q

What 3 conditions must our tissue samples be kept under, when carrying out cell fractionation

A
  • Buffered
  • Isotonic
  • Cold
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4
Q

Why are tissue samples stored in a buffered solution

A
  • Prevents PH changes, so that hydrogen and ionic bonds in the tertiary structure of proteins within organelles, remain unaffected
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5
Q

Why are the tissue samples stored in an isotonic solution

A
  • Equal water potential, therefore water doesn’t move into organelles via osmosis, preventing the lysis of organelles
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6
Q

Why are tissue samples stored in a cold solution

A
  • Reduces enzyme activity, which prevents the hydrolysis of proteins within organelles
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7
Q

Describe how cells can be homogenised

A
  • Physically using a mortar and pestle
  • Chemically using a weak detergent
  • The cell membranes are disrupted, to release the organelles within
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8
Q

Describe what happens after homogenisation

A

The solution is filtered to remove unbroken cells and large pieces of debris, forming a mixture called a homogenate

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9
Q

Describe what happens in centrifugation

A
  • A centrifuge separates things according to their different densities
  • The homogenate is placed in a centrifuge and spun
  • The most dense organelle falls to the bottom (pellet) and is removed, and the liquid on top (supernatant) will be spun at a faster speed to cause the next most dense organelle to fall to the bottom
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10
Q

Starting with some lettuce leaves, describe
how you would obtain a sample of undamaged
chloroplasts. Use your knowledge of cell
fractionation and ultracentrifugation to
answer this question.

A
  • Chop up (accept any reference to crude breaking up);
  • Cold;
  • Buffer solution;
  • Isotonic / same water potential;
  • Filter and centrifuge filtrate;
  • Centrifuge supernatant at higher speed;
  • Chloroplasts in (second) pellet;
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