Exam 1: Western Blot Part II

Question 1

Why is Western Blot more specific than the Lab 4 Experiments with fish?

Answer 1

• it uses antibodies

Question 1

Goals of Western Blotting

Answer 1

• determine presence of specific proteins
• quantify expression levels
• determine whether proteins have undergone genetic or posttranslational modifications
• identification of proteins based on molecular mass and antibody binding specificity

Question 2

Western Blotting

Answer 2

• also called immunoblotting
• antibodies are produced by immunizing animals with specific protein and collecting the antibodies that are produced

Question 3

Antibodies

Answer 3

-key proteins found in immune system, used to detect foreign invaders such as viruses bacteria, allergens
- specific and unique antibodies generated against invaders
- have ability to attach to specific target or epitopes of protein, thus making it possible to be used for immunodetection

Question 4

Review Image of Antibody vs Antigen

Answer 4

Notice structural difference in viewing just images

Question 5

Review manufacturing antibodies image

Answer 5

Note the difference between polyclonal and monoclonal

Note process and how to explain it

Question 6

HRP

Answer 6

enzyme on secondary antibody that catalyzes oxidation of the colorimetric substrate so the protein of interest can be identified

when adding colorimetric enzyme to substrate to blot, purple/gray bands will develop