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Questions and Answers
What is the primary purpose of homogenization in the context of protein extraction?
What is the primary purpose of homogenization in the context of protein extraction?
Which technique is employed immediately after cells have been ruptured to help separate different components?
Which technique is employed immediately after cells have been ruptured to help separate different components?
Which statement is NOT true regarding the Potter-Elvehjem homogenizer?
Which statement is NOT true regarding the Potter-Elvehjem homogenizer?
What alternative method can be used to rupture cells, as mentioned in the content?
What alternative method can be used to rupture cells, as mentioned in the content?
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How does differential centrifugation assist in the purification of proteins?
How does differential centrifugation assist in the purification of proteins?
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Study Notes
Protein Purification and Characterization Techniques
- Many different proteins exist within one cell.
- Many techniques are used to remove contaminants and isolate pure protein samples.
- The first step is releasing proteins from cells and organelles.
- Homogenization breaks open cells.
Homogenization Techniques
- Potter-Elvehjem homogenizer: A thick-walled test tube with a tight-fitting plunger is used to break open cells. The tube is moved up and down to grind the tissues.
- Blending: Tissues are ground in a blender with a suitable buffer solution.
- Sonication: Sound waves are used to break open cells.
Differential Centrifugation
- To separate particles, ruptured cells are centrifuged multiple times.
- Increased gravitational force at each step allows for separation of different-sized particles, which form pellets of different cellular components at various spin speeds.
- Pellet 1: Nuclei and unbroken cells
- Pellet 2: Mitochondria, Lysosomes, Microbodies
- Pellet 3: Ribosomes, Microsomes (ER, Golgi, plasma-membrane fragments)
- Supernatant : The liquid part remaining after centrifugation
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Description
Explore the various methods used for protein purification and characterization, including homogenization and differential centrifugation. This quiz will test your understanding of different techniques employed to isolate proteins and remove contaminants in a laboratory setting.
