Protein Purification Techniques
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Questions and Answers

What is the first step in isolating proteins from cells?

Release protein from cells and organelles

What technique is used to break open cells?

Homogenization

What is the purpose of straining the homogenate?

Remove extracellular material

What is the purpose of differential centrifugation?

<p>Separate particles based on their size and density</p> Signup and view all the answers

Which of the following is NOT a technique used to homogenize cells?

<p>Microscopy (D)</p> Signup and view all the answers

The force of gravity used in differential centrifugation remains constant throughout the process.

<p>False (B)</p> Signup and view all the answers

What is the name of the technique used to separate particles based on their size and density?

<p>Differential centrifugation</p> Signup and view all the answers

What is the ultimate goal of protein purification?

<p>Obtain a pure sample of the protein of interest</p> Signup and view all the answers

Flashcards

Protein Purification

The process of isolating a specific protein from a mixture of proteins, often found in cells.

Cellular Protein Extraction

The first step in protein purification, involving releasing proteins from cells and organelles for further analysis.

Homogenization

A technique used to break open cells and release their contents for protein extraction. It involves disrupting the cell membrane.

Potter-Elvehjem Homogenizer

A tool used for homogenization, consisting of a thick-walled test tube with a plunger that disrupts cells by pressing against the tube wall.

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Differential Centrifugation

A method of separating cellular components by spinning them at different speeds. Heavier components settle first due to gravity.

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Extracellular Material

Substances found outside the cells, like cell debris or other molecules, that need to be removed during protein purification.

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Sonication

A method of homogenizing cells by using high-frequency sound waves that create bubbles that disrupt cells.

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Grind Tissue in a Blender

A crude method to homogenize cells, where tissue is blended with a suitable buffer to break down cells.

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Purification

The process of separating and isolating the protein of interest from other cellular components.

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Contaminants

Unwanted substances that need to be removed during protein purification, such as other proteins or cellular debris.

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Pure Sample

A sample containing only the intended protein, with all other contaminants removed.

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Organelles

Specialized compartments within cells that have specific functions, like mitochondria or the nucleus.

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Cell Membrane

The outer boundary of a cell, separating its contents from the external environment.

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Centrifuge

A machine that spins samples at high speeds, separating components based on their density.

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Force of Gravity

The force that pulls objects towards the center of the Earth, used in centrifugation to separate components.

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Spin Speeds

The speed at which the centrifuge spins, affecting the separation of different components.

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Particle Separation

The process of separating different cellular components into distinct layers based on their density during centrifugation.

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Suitable Buffer

A solution with specific pH and ionic strength, used in homogenization to maintain protein stability.

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Study Notes

Protein Purification and Characterization Techniques

  • Cells contain various proteins
  • Techniques isolate pure protein samples
  • First step: release proteins from cells and organelles
  • Homogenization breaks open cells

Homogenization Methods

  • Potter-Elvehjem homogenizer:
    • Thick-walled test tube
    • Plunger passed through tube
    • Breaks open cells
    • Removes extracellular material by straining
  • Blender:
    • Grind tissue with suitable buffer
  • Sonication:
    • Sound waves break open cells

Differential Centrifugation

  • Ruptured cells centrifuged repeatedly
  • Increasing force of gravity each time
  • Differential spin speeds separate particles based on size and density
  • Different pellets are formed at different stages of centrifugation
  • Homogenate spun at low speed, and supernatant is collected.
  • Subsequent supernatants are spun at increasingly higher speeds to separate progressively smaller components

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Related Documents

Protein Purification and Characterization Techniques PDF

Description

This quiz covers the essential techniques for protein purification and characterization, including homogenization methods and differential centrifugation. Learn about various methods such as the Potter-Elvehjem homogenizer, blenders, and sonication, along with the principles behind differential centrifugation for isolating protein samples.

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