TESTS

Question 1

Edman Degradation

Answer 1

Identifies sequences in N terminus up to 15 residues

Enough to ID protein, but not efficient.

Question 2

Mass spectrometry

Answer 2

Determines precise molecule mass

Can detect minor species

Question 3

Proteolysis

Answer 3

Cleaves small peptides with pro teases.

Uses chymotripsin and trypsin to generate specific fragments

Fragments are analyzed with mass spec or electrophoreisis. Cleaves specific fragments and is useful for that.

Question 4

X Ray Crystallography

Answer 4

Refraction pattern that is shown reflects the 3D structure of the protein. Does not ID protein that is unknown.

Question 5

NMR

Answer 5

Determines structures of small protein in solution.

3D structure will not ID the protein

Question 6

Electrophoreisis- native

Answer 6

Separates molecules by charge, shape, size.

Non Denaturing. The rate of migration in gel IDs the protein.

Question 7

Electrophoreisis - SDS

Answer 7

Breaks down protein backbone. SDS binds to the backbone, making mass:charge, so that migration is a reflection of mass.

Determines mass, and number of subunits (bc of breakdown)

Question 8

ELISA

Answer 8

Highly sensitive means of detecting protein from a SMALL sample, using ANTIBODY.

Direct or indirect- whether or not a second antibody is required for tagging.

Question 9

Western Blotting

Answer 9

Separates protein from mixture by size and charge, and then by match to an ANTIBODY

Antibody used is specific to the protein in question - but have to know what you’re looking for, so it is more of a detection technique.

Can be used on proteins that coexist (copreciptiation)

Question 10

Immunoaffinity Chromatography

Answer 10

Column of ANTIBODY used to isolate, then elute the protein of interest.

Separates the protein, can can be used on coexisting proteins.

Question 11

Immunohistochemistry

Answer 11

Blood/cell sample used

Antibody is added to the sample, and then a second antibody with indicator.

Shows specific proteins in he tissue with staining.

Question 12

Epitope Tagging

Answer 12

Used when good antibodies for the protein can’t be developed.

Detects recombinant proteins

Epitope=tag

Useful in lab, not in clinical setting.

Question 13

Sickle Cell test

Answer 13

Electrophoreisis detects HbS.

Question 14

Doping

Answer 14

Electrophoreisis detects erythropoietin type.

Question 15

HIV

Answer 15

ELISA, followed by Western Blotting to confirm

Question 16

Myocardial Infarction

Answer 16

ELISA detects presence of MI indicator troponin

Question 17

P53 and MDM2

Answer 17

Western Blotting copreciptiation

Question 18

HER2

Answer 18

Immunohistochemistry