Antibody

Front 1

What is an antibody?

Back 1

Antibody or immunoglobulin is a specialized glycoprotein, produced from activated B cells (plasma cells) in response to an antigen

Front 2

Ig constitutes ____% of total serum proteins

Back 2

20-25%

Front 3

Electrophoresis of human serum proteins

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Front 4

General structure of antibody

Back 4

Front 5

H chains

Back 5

γ, α, µ, δ and ε

Front 6

In humans, __ of the light chains are kappa and ___are lambda type (ratio 3:2)

Back 6

60%

40%

Front 7

hypervariable regions or complementarity determining regions (CDRs)

Back 7

Within the variable region, there are some zones (hot spots) that show relatively higher variability in the amino acid sequences

There are three hot spots in the L and four in the H chain, respectively

Front 8

Features of hinge region

Back 8

This region is rich in proline and cysteine. The hinge region is flexible.

The hinge region is sensitive to various enzymatic digestions.

Front 9

Enzymatic digestion of immunoglobulin

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Front 10

the binding of an antibody to its antigen does not result in any direct biological effect. Rather, variety of secondary “effector functions” are produced

These effector functions include:

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1.Fixation of complement

2.Binding to various cell types

Phagocytes, lymphocytes, platelets, mast cells, NK cell, eosinophils and basophils

placental trophoblasts(IgG)

Front 11

IgG constitutes about ____ of total Ig in the body.

Back 11

70-80%

Front 12

What is the half life of IgG and how many subclasses does it have?

Back 12

23 days

4 subclasses

Front 13

complement-fixing ability of subclasses

IgG1 , IgG2 , IgG3 , IgG4

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IgG3 > IgG1 > IgG2

IgG does not fix complements

Front 14

Which subclass of IgG is involved in phagocytosis?

Back 14

IgG1 and IgG3 bind to Fc receptors present on phagocytes (macrophages, neutrophils) with high affinity and enhance the phagocytosis (opsonization) of antigen bound to them. IgG2 has an extremely low affinity for Fc receptors of phagocytes

Front 15

Why does IgG play a major role in neutralization of toxins ?

Back 15

as it can easily diffuse into extravascular space

Front 16

Which reactions does IgG mediate & what does it represent?

Back 16

•Precipitation & neutralization

•Chronic or past illnes(raised after a long time following infection)

Front 17

How does IgG mediate coagglutination?

Back 17

IgG subclasses (except IgG3) mediate coagglutination reaction by binding to protein-A of S. aureus

Front 18

Where is IgM present?

Back 18

present only in intravascular compartment, not in body fluids or secretions

Front 19

In how many forms IgM exists?

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1.Monomeric ( When present as membrane-bound antibody on B cells)

2.Pentameric (When present in secreted form)

Front 20

Overall functions of IgG

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1. Complement fixing

2. Phagocytosis

3. Coagglutination

Front 21

Functions of IgM

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1.Acute infection (first Ig to be produced following an infection)

2.complement fixing (most potent activator of classical complement pathway-5 Fc regions)

3.serves as B cell receptor for antigen binding

4.opsonin( IgM>IgG - 500 to 1000times)

5. Fetal immunity

6. Protection against intravascular organisms

7.Mediates agglutination (IgM>IgG - 20 times)

Front 22

What is to the % of IgA in total serum &

In what r the forms in which it exists?

Back 22

10 - 15%

•Monomeric (IgA in serum)

• dimeric ( secretory IgA)

- J chain , secretory component

Front 23

Functions of serum IgA

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initiates various functions such as •antibody-dependent cell-mediated cytotoxicity (ADCC),

•degranulation of immune cells etc.

Front 24

Where is secretory IgA predominantly found?

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body secretions like milk, saliva, tears, intestinal and respiratory tract mucosal secretions

Front 25

Functions of IgA

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mediates local or mucosal immunity

effective against bacteria like Salmonella, Vibrio, Neisseria &

viruses like polio and influenza

Front 26

Where r these synthesized

Dimeric secretory IgA

J chain

Secretory component

Back 26

Dimeric s.IgA & J chain - plasm cells situated near mucosal epithelium

Secretory component - mucosal epi.cells

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Function of secretory component of IgA

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1. Helps IgA cross epithelial surface & reach lumen

2. protects IgA from denaturation by bacterial proteases produced by intestinal flora.

Front 28

Subclasses of IgA

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1. IgA1 ( serum)

2. IgA2 ( secretions)

Front 29

Features of IgE

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1. Has Lowest serum conc

2.shortest half life & daily production

3. Only heat labile Ig

4. Homocytotrophism

5. Mainly extravascular in distribution

Front 30

Functions of IgE

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1. Mediates type I hypersensitivity rxns

(by binding to the mast cells causing degranulation.) Seen in allergies

2. Elevated in helminthic infections

( Coats on the surface of eosinophils

ADCC)

Front 31

IgD

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Front 32

On what basis Ig molecules are divided into isotypes, idiotypes and allotypes?

Back 32

Based on the location of antigenic determinants

Front 33

Antigenic determinants of immunoglobulin

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Front 34

How do isotypes vary from each other?

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in the amino acid sequences of the constant region of their heavy chains.

Front 35

What are idiotopes and idiotypes?

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The unique amino acid sequence present in paratope region (in VH and VL regions) of one member of a species acts as antigenic determinant to other members of the same species. Such antigenic determinants are called as idiotopes and the sum total of idiotopes on an Ig molecule constitutes its idiotypes

Front 36

Does somatic hypermutations in idiotypes evoke AI response

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They may act as foreign to the host itself but are present in small numbers .So do not evoke AI

Front 37

What are allotypes immunoglobulins

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The antigenic determinants present in the isotype genes in the constant region of H and L chains, encoded by multiple alleles are called as allotypes.

Front 38

When are abnormal immunoglobulins like Bence jones proteins produced?

Back 38

produced in a neoplastic condition of plasma cells called multiple myeloma / light chain disease which are accumulated in patient’s serum and excreted in urine

•coagulation at 50° & redissolving at 70°

Front 39

What are the other abnormal immunoglobulins?

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1.Waldenstrom’s Macroglobulinemia

(B cell lymphoma - excess IgM)

2.Heavy chain disease(exssve heavy ch)

3.Cryoglobulinemia( seen in Multiple mye & hep C infection)

Front 40

What are monoclonal antibodies?

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The antibodies derived from a single clone of plasma cell; all having the same antigen specificity, i.e. produced against a single epitope of an antigen.

Front 41

How are monoclonal antibodies produced & who developed it?

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Monoclonal antibodies are produced by Hybridoma technique, developed by G Kohler and C Milstein

Front 42

When are polyclonal & monoclonal antibodies produced in serum?

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•When an antigen having multiple epitopes enters the body -POLY

•when only one clone of B cell is stimulated by a single epitope of an antigen - MONOCLONAL

Front 43

What is the principle of hybridoma technique?

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Hybridoma technology

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Front 45

Steps of hybridoma technique

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1.Mouse splenic Bcell

2.Myeloma cells(double mutated)

3.Fusion(3 types generated)

4.purification

5.HAT medium(aminopterin×de novo)

•hypoxanthine, aminopterin,thymidine

-salvage pw(2 enzymesHGPRT&TKinase)

-myeloma lacks HGPRT

6.Fate of three types of cells on HAT media

7.Selection of individual hybridoma cells

-diluted; selected by RIA/ELISA

8.Maintainance of mAb(cultured-pure)

(Ascitic fluid -mixed with other Ab)

•purifican- chromatography

Front 46

What is the concentration of hybridom cells produced in pure form & mixed form

Back 46

Pure form- 10-60 ug/ml

Mixed form - 1-10mg/ml

Front 47

What happens if we inject mouse derived mAb into humans?

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the mouse proteins being foreign; can induce immune response in humans producing

human anti-mouse antibodies (HAMA)

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Types of mAb

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What are the postulates which explain the mechanism of antibody diversity?

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1. Multiple chromosomes

2. Multiple genes exist for each segment