Antigen Structure and Immunogenicity Updated: September 22, 2015 Folder Title: Antigens (With TP...

Antigen Structure and Immunogenicity

Updated: September 22, 2015

Folder Title: Antigens(With TP Slides)

Antigen Recognition by the Specific Immune System:Recognition by Antibodies on Bone-marrow Derived Lymphocytes (B-Cells)

or by Antibodies Secreted by B-Cells (Plasma Cells)or

Recognition by T-Cell Receptors on T-Cells (Thymus-derived Lymphocytes)

Antigen (Ag): Binds specifically to an antibody binding site (Ab), or to a T-cell receptor (TCR)*

• (* When the antigen is being presented to the TCR on a specific set of cell-membrane proteins called Major Histocompatibility Proteins (MHC)

Immunogen: Binds specifically to an antibody binding site or to a T-cell receptor*,

and generates a humoral or cellular immune response.

All immunogens must be antigens, Not all antigens can generate a response.

Haptens and Epitopes Haptens: Small organic molecule

(e.g. 1-Fluoro-2,4-dinitrobenzene)

Not Immunogenic by themselves

Can be immunogenic when attached to an immunogenic macromolecular carrier

The Anti-hapten antibody so generated will bind the Hapten

Epitope: A region of a macromolecular antigen that is actually an antigenic determinant,

a molecular sub-region that is actually recognized and bound by an Antibody or T-Cell Receptor

Fig.4-6bKuby3rd Ed

AgAb

Sep

Influenza Virus AntigenAntibody Heavy ChainVariable Region

Antibody Light Chain Variable Region

Antigen-Antibody Complementarity

Antigen - Antibody BindingAntibody Light Chain

Variable Region

Fig.

4-6a

Kuby

3rd Ed

AgAb

KissInfluenza Virus Antigen

Antibody Heavy Chain

Variable Region

Prot2nd

Primary sequential epitopes in

sperm whale myoglobin

(Epitopes are antigenic determinants inMacromolecules)

Conformational (non-sequential) Epitope in Hen

Egg White Lysozyme.

(Colors show amino acid side chains

contacting the two different chains of the

antibody, or contacting both

chains. Antibody structure to be covered later)

Myoglobin Single Polypeptide

17,000 MW (Tertiary Structure)

About 7 secondary domains. One bound Heme in hydrophobic

pocket

Hemoglobin: a2b2 Tetramer64,000 MWOne heme per subunit

“Fill-in-the Blank” Question:When a small organic molecule is made immunogenic

by attaching it to a macromolecule it is called a __ __ __ __ __ __

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What is this molecular

model showing?

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What Determines Immunogenicity of an Antigen?

1.Properties of the immunogen(See detailed breakdown next slide)

2.Properties of the Host (Slides 12 and 18)

a. Geneticb. Physiological

3. Mode of Antigen Exposure

ImGenFx1

ImGenFx2

Called “H2” in Mice; “HLA” in HumanHLA = Human Leucocyte-associated Antigens

Immunogenicity of Antigens and the MHC Gene Collection:MHC = Major Histocompatibility ComplexHaplotype = a set of closely linked genes controlling the immune response. Haplotype is usually transferred to progeny as a linked set

Antigen #1 Antigen #2

Ag&MHCbAg 1 Ag 2

Two different antigens

Ag&MHCcAg 1 Ag 2

Two different antigens

Ag&MHCdAg 1 Ag 2

Two different antigens

Ag&MHCAg 1 Ag 2

Two different antigens

ImGenFx3

ImAdmin

To here: Tuesday,Sept. 15, 2015

Conformational Properties of Epitopes for Antibodies

64

80

Hen Egg-White Lysozyme: Conformations of Sequence 62 to 80

Anti-hapten Antibodies

AbHaptenSee Figure 4-1p. 77Kuby, 6th Edition

AgByB&Ta

AgByB&Tb

AgByB&T

Conformational (non-sequential) Epitope in

Hen Egg White Lysozyme.

To here: Thursday,Sept. 17, 2015

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This graph shows two forms of the amino acid sequence between cysteine 64 and cysteine 80 in hen egg white lysozyme

(HEL). One form of the synthetic antigenic region does not inhibit the

normal reaction between HEL and its antibody while the other form does inhibit. Why does the form of the

potential antigenic sequence shown in green fail to inhibit the antigen-antibody reaction between HEL and its antibody?

What does this table tell you about the specificity of antibody recognition for antigens?

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To here: Thursday,Sept. 17, 2015

Antigens in Immunologically-Based Clinical Diagnosis and Pathology

Cancer Antigens in Diagnosis of Cancer

Carcino-Embryonic Antigen (CEA)

Prostate Specific Antigen (PSA)

Ovarian Cancer Antigen (CA 125)

Whole-body Imaging of Metastatic Colon Carcinoma

A33 Antigen

Anti-Human Chorionic Gonadotropin Pregnancy Test(Early Home Pregnancy Test)

Type II Allergic Hypersensitivity

Blood Group Antigens in Transfusions

Hemolytic Disease of New-Borns: Rh Antigen incompatibility

Super-Antigens in Toxic Shock Syndrome

Antigens in Immunologically-BasedTreatment of Cancers and Other Pathologies:

Cancer Antigens on Cancer Cells

Cancer Antigens as Targets for Therapy

Antibodies as Anti-cancer AgentsRadioactively-labeled Antibodies

Cytolytic Antibodies

Specifically-activated Cytotoxic T-Cells as Anti-Cancer Agents

Specifically Activated Antigen-presenting Cells (Tc).

Imaging with Antibodies Carrying “Reporter Groups”

Radioactive IsotopesFluorescent Groups (“Chromophores”)Metals

Antigen-Antibody Interactions in Research and Clinical Medicine

e.g. Radioactively labeled monoclonal antibody reacting with tumor cell antigens in detection of metastatic colon cancer

SeeMets

Arm

Head

Imaging on Metastatic Colon Carcinoma with Radioactive-Iodine-Labelled Monoclonal Ab to A33 AgLloyd Old, Scientific American, August, 1996, p. 138)

Antihapten Antibodies in the Detection and Measurement of Small Molecules

For example: Anti-HCG (Human chorionic gonadotrophin) antibody in the detection of soluble HCG in urine

AntiHCGWhat will happen when concentration of Anti-HCG-Ab is about equal to Hapten-carrier conjugate?

What will happen when you put excess soluble hapten in the way of the anti-HCG-Ab?

PregTest

Antibodies in Blood Transfusions

Antibodies to A, B, O Blood Group Antigens

Pre-existing Antibodies to Non-tolerated (Unmatched) Blood Group Antigens : “Isohemagglutinins”

Why are these non-matched Blood-group Antigens generating antibodies (Isohemagglutins) at all?

Because infectious bacteria do possess and exhibit cross-reacting antigens similar to blood group antigens

ABO Types

Pre-existing serum antibodies from exposure to bacterial antigens that mimic antigens on red blood cells

ABOSugar

See Figure 15-13 (a), p. 389,

Kuby, 6th EditionGalactose

Not a hexasaccharide(No terminal sugar)

Antibodies in Hemolytic Disease of the New-born (HDNB)

(Rh-Factor Sensitization)

Rh-Factor positive baby – Rh Negative Mother

HDNB

See Figure 15-14p. 390, Kuby6th Edition

HDNBPhoto

AntiRhAb

Following are three Turning Point short answer questions.

Please put all notes on the floor.Do not have any electronic devices other than your NXT transmitter.No consulting with other students.

If you have a problem with your device, I can provide you with a loaner NXT device.

If you have a problem using your NXT device, please ask Elisabeth for help.

It is imperative that the integrity of these in-class Turning Point quizzes be maintained at the same level as we will do with the three written exams.

“Fill-in-the Blank” Question:Hemolytic disease of the new-born is caused by maternal

__ __ __ __ __ __ __ __ __ __attacking the fetus in utero.

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“Fill-in-the Blank” Question:What are those maternal antibodies recognizing and reacting against?

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“Fill-in-the Blank” Question:What is this Whole-body X-ray Showing?

Activation of Systemic Inflammatory Responses by Potent Inflammatory Signals

from Infectious Organisms and Plants

EndotoxinsExotoxinsSuper-antigensMitogens and Lectins

Immunopathology of Bacterial Septic Shock Caused by Gram Negative Bacterial Endotoxins:Examples -E. coliPseudomonas aeruginosaNeisseria meningitidisMeningococcus

70,000 Deaths per yearDiarrhea, Fever, Blood Clotting, Blood Pressure Drop

Macrophage Activation by Endotoxin:Generates systemic cytokine production -IL1 and TNF-alpha (Tumor necrosis factor)

Immunopathology of Bacterial Toxic Shock Exotoxins Secreted or Presented Membrane Bound:Act as "Superantigens"Examples -Staphylococcus aureus: TSST1 (Toxic Shock Syndrome Toxin)Streptococcus pyrogenes - Rheumatic fever and shockStaphylococcus enterotoxins: Food poisoning

Superantigens cross-link TCR with Antigen-presenting Cell MHC Class II Proteins

Generates systemic cytokine production - IL1 and TNF-alpha (Tumor necrosis factor)

SuperAg

See Figure 11-67th Edition p. 367

Staphylococcal enterotoxins (food poisoning)Streptococcal pyrogenes exotoxins (rheumatic fever, toxic shock)See Table 11-2, p. 367, 7th Edition

Mitogens and Lectins

Mitogens: Stimulate mitosis and cell division non-specifically(non-specific therefore stimulate polyclonally)

Lectins: Proteins that bind to carbohydrate in glycoproteins(Can be potent mitogens)

Glycoproteins: Conjugated protein with covalently attached carbohydrate residues

Examples of Mitogens

Concanavalin A (Con A) : Non-specific T-cell mitogen

Phytohemagglutinin (PHA): Non-specific T-cell mitogen

Pokeweed Mitogen (PWM): Non-specific T- and B-cell Mitogen

Lipopolysaccharide (LPS): Gram negative bacterial cell wall component

Potent Polyclonal (non-specific) B-Cell Mitogen

DescriptionGibco® Phytohemagglutinin, M form (PHA-M) is a solution made from a crude extract of the red kidney bean Phaseolus vulgaris. This extract is capable of inducing blastogenesis in vitro in various mammalian mononuclear cells when used at a concentration of 1–2% v/v. Each lot of Gibco® Phytohemagglutinin is performance tested at 1.5% using peripheral blood lymphocytes from a normal adult donor cultured for 72 hours in Gibco® RPMI 1640 Medium supplemented with Fetal Bovine Serum (FBS), heparin, antibiotics, and L-glutamine.

Google Description of Phytohemagglutin:What BIO 447 Is Intended to Achieve

Immune Adjuvants and the Immunogenicity of Vaccines(See Scientific American, October, 2009)

Boosting Vaccine Power, pp 72 to 79

Activating Antigen-presentation by Cells of the Innate Natural Immune Response

e.g. Purified derivatives of bacterial lipo-polysaccharidesBacterial DNA derivativesBacterial flagellin protein

Specific Activation of Antigen-presenting Cells Extra-corporeally (outside of the body)

Dendritic Cell VaccinesSee Dendreon

Innate Natural Immunity and Systemic Sepsis:NOVA Video on Meningitis (Meningococcal

Meningitis, Endotoxin, and Systemic Sepsis)

http://www.pbs.org/wgbh/nova/meningitis/

Has video links to steps in meningococcal sepsis

“Amy’s Story”and related NOVA Stories

tpfondy.mysite.syr.edu

The Story of the Oakwood Children

See “Song of the Oakwood Children” as a self-running presentation

Following are two Turning Point short answer questions.

Please put all notes on the floor.Do not have any electronic devices other than your NXT transmitter.No consulting with other students.

If you have a problem with your device, I can provide you with a loaner NXT device.

If you have a problem using your NXT device, please ask Elisabeth for help.

It is imperative that the integrity of these in-class Turning Point quizzes be maintained at the same level as we will do with the three written exams.

What is a “Mitogen”?What Does a Mitogen Do in Cell Biology Terms?

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“Fill-in-the Blank” Question:RhoGam is an antibody that prevents sensitization of the Rh- mother

by cells from her fetus. Why does that work?

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Merck Introduction of Keytruda

For Malignant Melanoma

Sept. 6, 2014Wall-Street Journal

Working Link Sent to Each Student by Yifan

Discussion and Comments

Whooping Cough Infection and Transmission

Whooping Cough VaccineToday Show, 12/2/04

To Here September 16th; Showed 6 minute video of whooping cough disease and vaccinations

Dendreon is a Commercial Vaccine that Uses Dendritic Cells to Generate an Immune Response. What is it that the Dendritic Cells do in Immunology that allow them to be used effectively in this way?

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End of Presentation on Antigens: Sept. 18, 2014

Save Session

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3. I’m following OK, I can catch up with what I miss.4. I’m struggling to keep up. It’s too much too fast.

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We watched parts of a video on Meningitis and Meningococcal Sepsis caused by an Infectious Bacterium. Killing the bacterium doesn’t immediately cure the sepsis. Why is that? What is causing the systemic sepsis?

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“Fill-in-the Blank” Question:The antibody to Human Chorionic Gonadotrophin fails to cross link the

HCG-hapten carrier conjugate and fails to make a precipitate if urine from a pregnant woman is added to the mixture. What is in the urine that

is preventing the formation of the precipitate? Abbreviations are OK______________________________________

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“Fill-in-the Blank” Question:Human chorionic gonadotrophin (HCG) is a small peptide made

immunogenic by being conjugated with a large molecular weight carrier molecule. Thus HCG is a special kind of antigen and is an example of a

__ ___ ___ ___ ___ ___

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“Fill-in-the Blank” Question:A small organic molecule such as fluoro-dinitro benzene

can be made immunogenic by attaching it to a _____________________________

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Immunogenicity of Antigens is Controlled by the Nature of the Antigen, the Genetics of the Host, and the Mode of Immunogen Exposure.

Give an example of what is meant by “Mode of Immunogen or Antigen Exposure”

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A Specific target for an Antibody is an_ _ _ _ _ _ _.

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