Separating Cell Structures By Fractionation

Source: Urry, Lisa A.. Campbell Biology (p. 96). Pearson Education. Kindle Edition.

A useful technique for studying cell structure and function is cell fractionation, which takes cells apart and separates major organelles and other subcellular structures from one another. The piece of equipment that is used for this task is the centrifuge, which spins test tubes holding mixtures of disrupted cells at a series of increasing speeds. At each speed, the resulting force causes a subset of the cell components to settle to the bottom of the tube, forming a pellet. At lower speeds, the pellet consists of larger components, and higher speeds result in a pellet with smaller components. Cell fractionation enables researchers to prepare specific cell components in bulk and identify their functions, a task not usually possible with intact cells. For example, on one of the cell fractions, biochemical tests showed the presence of enzymes involved in cellular respiration, while electron microscopy revealed large numbers of the organelles called mitochondria. Together, these data helped biologists determine that mitochondria are the sites of cellular respiration. Biochemistry and cytology thus complement each other in correlating cell function with structure.

Technique: Cells are homogenized in a blender to break them up. The resulting mixture (homogenate) is centrifuged. The supernatant (the liquid above the pellet) is poured into another tube and centrifuged at a higher speed for a longer period. This process is repeated several times. This “differential centrifugation” results in a series of pellets, each containing different cell components.

Results: In early experiments, researchers used microscopy to identify the organelles in each pellet and biochemical methods to determine their metabolic functions. These identifications established a baseline for this method, enabling today’s researchers to know which cell fraction they should collect in order to isolate and study particular organelles.

Source:

Urry, Lisa A.. Campbell Biology (p. 96). Pearson Education. Kindle Edition.

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