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MICR 201 Microbiology for Health Related Sciences

Lecture 11: Practical applications of immunology ; vaccinationsEdith Porter, M.D.

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Lecture outline

Major applications of immunology▪ Vaccines▪ Immunological memory▪ Types of adaptive immunity▪ Types of vaccines▪ Development of new vaccines▪ Safety of vaccines

▪ Generation of antibodies for research and diagnostic▪ Anti-sera▪ Monoclonal antibodies

▪ Diagnostic immunology▪ Precipitation and agglutination reactions▪ Complement fixation▪ ELISA, western blot, immunofluorescence

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Immunological memory

Once lymphocytes have encountered their specific antigen they undergo clonal expansion

Some of these cells develop further into memory cells Can circulate for many years

Upon re-contact with the same antigen they quickly proliferate and resume effector function B cells: antibody production T cells: cytokine production (TH, Treg) and cytotoxicity (CTL)

Instead of requiring 10 – 14 days for a full response measurable effects appear within 2 days

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Example: Primary and secondary immune responses to an antigen

IgM is always the first antibody

IgG follows IgM IgG level does not go back to

baseline Re-exposure to the same

antigen will lead to an augmented and accelerated immune response, increased IgG response and with higher residual antibody levels

Presence of antibodies is not equivalent to sickness

Primary Response

Secondary Response

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Types of adaptive immunity

Vaccines do not always target B cells and antibodies but some target T cells.

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Adaptive Immunity

Naturally acquired

Active immunity Resulting

from infection

Passive immunity Transplacent

al Colostrum

Artificially acquired

Active immunity Injection of Ag

(vaccination) Passive

immunity Injection of Ab

Vaccines: toxoid, killed microorganisms, live attenuated microorganisms

Long Lasting

Short Lasting

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Check your understanding

1) What type of immunity results from vaccination?

A) Innate immunity B) Naturally acquired active immunity C) Naturally acquired passive

immunity D) Artificially acquired active

immunity E) Artificially acquired passive

immunity

3) What type of immunity results from recovery from mumps?

A) Innate immunity B) Naturally acquired active immunity C) Naturally acquired passive

immunity D) Artificially acquired active

immunity E) Artificially acquired passive

immunity

15) The antibodies found in mucus, saliva, and tears are

A) IgG. B) IgM. C) IgA. D) IgD. E) IgE.

26) The best definition antibody is A) A serum protein. B) A protein that inactivates or kills an

antigen. C) A protein made in response to an

antigen that can combine with that antigen.

D) An immunoglobulin. E) A protein that combines with a

protein or carbohydrate.

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Principles of Vaccination

Induce active immune response with a related or inactivated agent that does not cause major disease

Induce cross-reactive antibodies or T cells Herd immunity is sufficient to prevent

epidemic diseases Must weigh the benefit of protection

versus the danger of vaccine associated disease

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Types of Vaccines (1)

Attenuated whole agent vaccines Live, weakened microbes Generated through long term culture Mimic most closely actual infection Caution: danger of backmutation to a virulent

form Dangerous for immunocompromised patients!

Inactivated whole agent vaccines Toxoid

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Types of Vaccines (2) Subunit or acellular vaccines

Use of antigenic subunit that triggers an immune response.

Inherent safer as it cannot reproduce in the host

Conjugated vaccine Combine as antigen poorly immunogenic

material (e.g. capsules) with an immunogenic protein

Nucleic acid vaccines (DNA vaccines) Introduce genes for protein targets into the

host Host will express the protein and mount an

immune response to this foreign protein

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Where to get the vaccine from?

Classical Bacterial culture (Animal extract) Cell culture Embryonated eggs

New Developments Recombinant vaccines Plants Influenza virus grown in

embryonated eggs

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Adjuvants

Improve effectiveness of vaccines/immunizations by providing co-stimulatory signals for T and B cells In humans▪ Alum▪ Oil based substance MF59 and virosomes

In animals▪ Freund’s complete adjuvants▪ Mycobacterium extract

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Diphtheria: Purified diphtheria toxoid Pertussis: Acellular fragments of B. pertussis

or antigenic acellular fragments Tetanus: Purified tetanus toxoid Meningococcal meningitis: Purified

polysaccharide from N. meningitidis Haemophilus influenzae type b meningitis:

Polysaccharides conjugated with protein Pneumococcal conjugate vaccine: S.

pneumoniae antigens conjugated with protein

Bacterial vaccines used in the US

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Influenza: Inactivated or attenuated virus Measles: Attenuated virus Mumps: Attenuated virus Rubella: Attenuated virus Chickenpox: Attenuated virus Poliomyelitis: Inactivated virus Hepatitis B: Antigenic fragments (recombinant) Smallpox: Live vaccinia virus Rabies: Inactivated virus Hepatitis A: Inactivated virus Human papilloma virus: Antigenic fragments

Viral vaccines used in the US

Selected patients

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Schedule of childhood immunizations

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Booster

Re-administration of vaccine to boost immune defense ( ) Increased antibody production Increased memory cell development

Time

Ab T

iter

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Success of vaccines

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Vaccines urgently needed

HIV Malaria Tuberculosis Cholera

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Concerns with vaccines

Safety Sometimes illness follows vaccination▪ Risk benefit analysis

Autism had been linked to vaccination▪ Most recent studies conclude that there is no

link but instead a genetic disposition Weaning protection

Bordetella pertussis

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Use of antibodies as a diagnostic Tool

Patients antibodies indicate that patient had contact with

the agent before “serology”

Commercial antibodies used to detect patient antigen

In research Detection of antigen Purification

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Serology Study of reactions between antibodies

and antigens Globulins

Serum proteins Gamma () globulin

Serum fraction containing Antibodies (immunoglobulin)

Antiserum Generic term for serum when it contains

specific Ab Polyclonal

Monoclonal antibodies

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Production of antiserum

Prepare antigen Inoculate animal with antigen and

adjuvants Administer several boosts Draw blood and let it coagulate Remove cells by centrifugation and

collect supernatant = serum with high titers of specific antibodies

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Production of monoclonal antibodies

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Selected serological tests

Precipitation Agglutination

Hemagglutination Fluorescent-antibody technique ELISA

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Precipitation reactions

Involve soluble antigens and antibodies

Upon cross linking a visible interlocking molecular aggregate is formed (lattice)

Only in equivalence zone precipitates are formed

Example Ouchterlony

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Precipitin ring test

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Agglutination reactions

Involve particulate antigens and antibodies

IgM are most efficient Direct agglutination

Antibodies against large cellular antigens

Indirect agglutination Antibodies against soluble

antigen adsorbed to a particle or erythrocytes

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Agglutination assay to measure antibody titer

Relates to the concentration of antibodies against a particular antigen

Gives a guide to how active the patient’s immune response is.

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Note: Viral hemagglutination is not an antigen:antibody reaction

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Viral hemagglutination inhibition for antibody detection

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Fluorescent antibody techniques (direct)

To detect difficult to grow pathogens (e.g. Legionella)

LabeledSpecific Antibody

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Fluorescent antibody techniques (indirect)

Figure 18.10b

SpecificPrimary Antibody

LabeledSecondary Antibody

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Enzyme-inked Immuno-Sorbent Assay (ELISA)

To detect Ag

To detect Ab

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Pregnancy test sticks: Mini ELISA:

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Important to remember

Vaccination (against disease) and immunizations (for antibody production) require administration of attenuated or inactivated antigen

Ab:Ag reaction is the fundamental reaction in immunology

Precipitation: ab--soluble ag Agglutination: ab--particulate ag Presence of antibodies does not mean

illness High titer means high concentration of

ab

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Check your understanding

4) In an agglutination test, eight serial dilutions to determine antibody titer were set up: tube #1 contained a 1:2 dilution; tube #2, a 1:4, etc. If tube #6 is the last tube showing agglutination, what is the antibody titer?

A) 6B) 1:6C) 64D) 1:32E) 32

16) A test used to identify antibodies against Treponema pallidum in a patient is the

A) Direct fluorescent-antibody test.B) Indirect fluorescent-antibody test.C) Direct agglutination test.D) Direct ELISA test.E) Hemagglutination-inhibition test.

15) What type of vaccine is live measles virus?

A) Conjugated vaccineB) Subunit vaccineC) Nucleic acid vaccineD) Attenuated whole-agent vaccineE) Toxoid vaccine

13) What type of vaccine involves host synthesis of viral antigens?

A) Conjugated vaccineB) Subunit vaccineC) Nucleic acid vaccineD) Attenuated whole-agent vaccineE) Toxoid vaccine