Separation Techniques 1.1

Question 1

what is a centrifuge used for?

Answer 1

to spin samples at high speeds to separate material according to density

Question 2

what are the high density materials that form at the bottom of the tube called?

Answer 2

a pellet

Question 3

what are the low density molecules that are formed on top of the tube called?

Answer 3

the supernatant

Question 4

what is chromatography used for?

Answer 4

to separate different substances such as amino acids and sugars

Question 5

what does the speed of each solute depend on?

Answer 5

it’s differing solubility in the solvent used

Question 6

describe paper chromatography

Answer 6

simple process involving a strip of chromatography paper

Question 7

describe thin layer chromatography

Answer 7

involves a thin layer of absorbent silica gel, alumina or cellulose over a slide of glass

Question 8

what are the benefits of thin layer chromatography

Answer 8

runs faster

better separations

given the choice of different absorbance

Question 9

what is affinity chromatography used for?

Answer 9

separating proteins

Question 10

what is gel electrophoresis used for?

Answer 10

separating proteins and nucleic acids

Question 11

how do native gels work?

Answer 11

separates proteins by shape, size and charge

does NOT denature the molecule

Question 12

how does SDS-PAGE work?

Answer 12

separates proteins by size alone

gives all molecules an equally negative charge and denatures them

Question 13

how can proteins be separated from a mixture?

Answer 13

by using their isoelectric points (IEPs)

Question 14

what happens if a solution is buffered to a specific pH?

Answer 14

only the protein(s) that have an IEP of that pH will precipitate

Question 15

how can proteins also be separated?

Answer 15

by using their IEPs in electrophoresis

Question 16

describe how affinity chromatography works

Answer 16

• solid matrix/gel is created with specific molecules (usually receptors) bound to matrix/gel

• soluble target proteins in a mixture with a high affinity for these molecules become attached as mixture passes down column

• non-target molecules are washed out due to low affinity

Question 17

describe how gel electrophoresis works

Answer 17

• charged macromolecules move through an electric field applied to a gel matrix

• samples are loaded into wells in a gel and an electric current is ran through it

• charged molecules will move towards the opposing charge
•smaller molecules will travel farther than larger molecules