1.1 Separation techniques

Question 1

Centrifuge

Answer 1

Substances are spun, more dense components settle in the pellet; less dense components remain in the supernatant.

Question 2

Paper and thin layer chromatography

Answer 2

Used for separating different substances such as amino acids and sugars. The speed that each solute travels along the chromatogram depends on its differing solubility in the solvent used.

Question 3

Affinity Chromatography

Answer 3

Used to separate proteins, A solid matrix or gel column column is created with specific molecules (usually receptors) bound to it. Soluble, target proteins in a mixture, with a high affinity for these molecules, become attached to them as the mixture passes down the column. Other non target molecules with a weaker affinity are washed out.

Question 4

Gel electrophoresis

Answer 4

Charged macromolecules move through an electric field applied to a gel matrix. Separating proteins and nucleic acid.

Question 5

Native gels

Answer 5

Separate proteins by shape, size and charge. And do not denature the molecules.

Question 6

SDS-PAGE

Answer 6

Separates proteins by size alone, gives all molecules an equally negative charge and denatures them, separating proteins by size alone.

Question 7

Protein separation by isoelectric points (IEP)

Answer 7

Proteins can be separated by their IEP, if the solution is buffered to a specific pH, only the proteins that have the IEP of that pH will precipitate.
The IEP is the pH at which soluble protein has no net charge and will precipitate out of solution.

Question 8

Proteins separation by IEP in electrophoresis.

Answer 8

Soluble proteins can be separated using an electric field and a pH gradient. A protein stops migrating through the gel at its IEP in the pH gradient because it has no net charge.