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Separation Methods Used in Biology Labs

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  • 0:04 Why Separate?
  • 0:44 Separation by Mass
  • 1:39 Separation by Affinity
  • 2:28 Electrophoresis Separation
  • 3:05 Multiple Parameter Separation
  • 3:51 Lesson Summary
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Lesson Transcript
Instructor: Meredith Mikell
Research in cell and molecular biology involves the separation of proteins and other cell components. Here we will investigate some of the key separation methods commonly used. At the end, you can test your knowledge with a brief quiz.

Why Separate?

Cells are complex little structures comprised of wildly different components that each serve specific functions. Proteins are one such component that are commonly studied for their importance in biological function. Biochemists and molecular biologists study the intricate roles of each protein and, in doing so, need to categorize them independently of each other. This is where separation methods come in handy. Proteins differ in their sizes, shapes, polarity, charges, and affinity to binding with other proteins. Based on these differences, proteins can be sorted and separated, much like sifting sand through a sieve and collecting the pieces of shell that don't make it through.

Separation by Mass

One of the most basic ways of separating cell components is to do so by mass. The size and weight of the organelles or other large particles can be sorted using a centrifuge, a device that spins cells at high speeds to generate substantial g-forces, or gravity due to centrifugal forces. Some centrifuges also implement heat or vacuum as a secondary means of separation. Cells or cell components are placed in a test tube containing a suspension liquid. After centrifugation, the largest components are gathered at the bottom of the tube, with progressively smaller components on top, comprising the precipitate, or solid portion of the solution. The remaining liquid, or supernate, is on top of the precipitate. Using a pipette, the precipitate can be removed and examined. This method is fairly simple and relatively inexpensive, though only effective at separating larger cell components.

Separation by Affinity

Many proteins have a particular affinity for different phases of substance mobility. In other words, they like to be stationary or they like to be on-the-go, based on how much they are attracted to other substances moving past them. A technique called chromatography utilizes this affinity to aid in the separation of proteins. There are several different types of chromatography that involve different ways that the proteins are attracted to other molecules. When the affinity is based on charge, ion exchange chromatography is used, wherein a charged gel or solution is used to attract and retain proteins of like charge. When the affinity is based on molecule size, gel filtration chromatography is used, wherein the protein solution is passed through gel beads that proteins of different size and affinity either pass through or around, thus separating.

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