Li - Antibodies

Question 1

Ways of generating an infinite amount of antibodies

Answer 1

1) different combos of heavy and light chains
2) combinatorial VDJ joining
3) Junctional diversity
4) alternative RNA splicing
5) class switch recombination
6) somatic hyper mutation
7) genetic variation

Question 2

On what chromosome(s) can you find the heavy chain?

Answer 2

14

Question 3

On what chromosome(s) can you find the light chain?

Answer 3

2, 22

• 2 - makes IGL-lambda
• 22 - makes IGL-kappa

Mnemonic - Leor Kushner, Leor is his first name so it is on chromosome 2 and Kushner is his last name so it is not chromosome 22.

Question 4

VDJ SOmatic Recmbination

Answer 4

Heavy chains - You have to select from 40 V genes, 27 D genes, 6 J genes, and 9 C genes, which makes for massive diversity.

Light chains - you have two different chromosomes with different amounts of V, J and C regions on them. For kappa you have 40 V, 5 J, and 1 C. For lambda you have 32 V and 4 J. - No for either

Question 5

Junctional Diversity

Answer 5

Following the gene sequence there are flanking RSS sequences. In the D region they are on both sides of each gene segment. In the V and J region they are only on one side. During recombination some of the RSS might be deleted, duplicated, mutated, etc. which adds the the diversity of antibodies.

Question 6

Class Switching Recombination

Answer 6

When you switch part of the constant region so that instead of being one type of antibody it becomes a different type of antibody. The variable region stays the same though so it still binds the same antigen.

Question 7

What role does Alternative RNA splicing play in antibody diversity?

Answer 7

Within the constant region you have have different parts being included into the final antibody (exons) from the transcript depending on how you cut it and which parts, which increases the diversity of antibody generation.
- If you think about it, the VDJ(C) part is just the DNA transcript, it can still be alternatively RNA spliced afterwards.

Question 8

SOmatic Hypermutation

Answer 8

Programmed mutation in the variable region of B cells in order to increase diversity.

Question 9

Genetic Variation

Answer 9

Mom and dad have different amounts of gene segments in the V, D, and J regions.
Within that there could Also be deletions/mutations in the chromosome.

Question 10

Monoclonal antibodies

Answer 10

Used as drugs. They are cloned from one B cell. Very specific for a specific antigen.
What they do is that they will inject a mouse with an antigen and then take out the Antibodies generated. They will mix these with myeloma cancer cells in order to make the cells immortal.
- The point of this is that you now have a single B cell that secrete single antibodies, and you know what it is acting on. SOmtimes, an antigen can have a lot of different antibodies. That act on different epitopes on it. A monoclonal antibody will recognize that one epitope on that one protein so it is extremely specific. Therefore, if there is something in the body that is not right, you can make an antibody against it.
- Anything that has “mab” in it as a monoclonal antibody.

Question 11

How are antibodies used in diagnostic testing?

Answer 11

ELISA

• Coat plate wells with specific Ab
• Add antigen to that antibody
• Add a detecting Ab to that antigen with biotin on the end of it.
• add horseradish per oxidase with streptavadin
• Add TMB to see the color.

Question 12

FcGR2B

Answer 12

This gene controls the production of IgG. If you produce too much of it you will have an autoimmune issue. If you produce too little you will be immunodeficient.

Question 13

Detailed structure of antibody

Answer 13

• composed of a light chain and a heavy chain
• the two heavy chains are connected by disulfide ones between CH1 and CH2
• The heavy and light chains are connected on each side by disulfide bonds
• The variable regions are on the N terminal side, the constant region is on the C terminal side
• COnstant region is split into three regions: CH1, CH2, CH3
• variable regions is split into 6 regions, the part that the epitope actually binds is called the CD-R region. There are also FR regions in the variable region.

Question 14

Paratope

Answer 14

The part of the antibody That actually binds to the epitope

Question 15

Where are B cells made?

Answer 15

In the bone marrow. As opposed to T cells, which are technically start in the bone marrow and then are really made in the thymus, B cells are made in the bone marrow completely.

Question 16

Germinal Center

Answer 16

This is called a germinal center of the spleen or lymph node. During an infection, the activated B cells travel to the dark zone in order to undergo clone lexpansion. Then from there, there are various mutations and added diversity that occurs in the light zone before various B cells are spit out.

Question 17

In a primary immune response vs. secondary immune response, what is the difference in what antibodies are secreted?

Answer 17

In a primary it is mostly IgM, in a secondary it is mostly IgG

Question 18

Mantle Cell Lymphoma

Answer 18

A cancer due to a translocation between chromosome 14 and 11 that puts Cyclin-D1 and IgH enhancer on the same chromosome, which causes upregulation of Cyclin-D1.

Question 19

Papain Cleavage

Answer 19

Produces two Fab fragments and one Fc fragment

Question 20

Fab

Answer 20

One entire light chain and the N terminal of the heavy chain. This allows binding to the antigen. There will be two of these in papain cleavage.

Question 21

Fac

Answer 21

Contains the C terminus ends of the heavy chains. It is responsible for many of the effector functions of the antibody like complement, binding to cell membrane, and placental transport. There is one of these in papain cleavage.

Question 22

Pepsin cleavage

Answer 22

Produces one Fab’2 molecule

Question 23

Fab2’

Answer 23

It is essentially two Fab molecules linked together at the hinge region. This is not done. Naturally and needs to be done in a lab.

Question 24

Idiotype

Answer 24

Unique structure of an antibody combining site, produced by V D and J segments combined with the various other mechanisms.

Question 25

12/23rule

Answer 25

The V, D, and J genes are separated by a 12 base spacer and a 23 base spacer, which is 1 or 2 turns of DNA.

Question 26

DNA Terminal Deoxyribonucleotidyl Transferase

Answer 26

It adds a random number of nucleotides after each segment of either V D or J in order to add diversity.

Question 27

The presence of which antibody is a sign of a recent NEW infection?

Answer 27

IgM. In a primary response, IgM is the antibody that is most common. In a secondary response we see IgG.