Cell fractionation

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8 Terms

1
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1st step

Cut tissue in ice cold isotonic buffer

Cold to slow enzyme reactions

Isotonic to stop osmosis, so organelles don’t burst

Buffer to stop pH changes

2
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2nd step

Grind tissue in a blender to break open cells

3
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3rd step

Filter. This removes insoluble tissue e.g. fat connective tissue

This filtrate is now called a cell free extract and is capable of carrying out most of the normal cell reactions

4
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4th step

Centrifuge filtrate at low speed

This pellets nuclei, which can be resuspended

5
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5th step

Centrifuge supernatant at medium speed

This pellets mitochondria and chloroplasts, which can be resuspended

6
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6th step

Centrifuge supernatant at high speed

This pellets ER, golgi and other membrane fragments which can be reususpended

7
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7th step

Centrifuge supernatant at very high speed

This pellets ribosomes which can be resuspended

8
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8th step

Supernatant is now organelle free cytoplasm