Ch 17: Vaccines and Detection

Immunization

Process of inducing immunity

Probs greatest impact on human health of any other medical procedure

Also immune reactions have led to useful diagnostic tests

Immunity can be

Naturally or artificially acquired

Naturally acquired

Gained through normal pathogen interaction (illness)

Artificially acquired immunity

Gained through vaccines

Active immunity

Results from exposure to antigen (lymphocytes activated)

Passive immunity

Results from addition of other’s antibodies (no memory activated)

Ex of natural active immunity

Exposure to infectious agent (pathogen)

Artificial active immunity ex

Vaccination

Natural passive immunity

Mother to child (during pregnancy)

Artificial passive immunity

Antibodies from other’s blood

Vaccine

Any preparation of a pathogen or its products used to induce active immunity

Protects individual; can prevent spread of pathogen in population

Herd immunity

Develops when a critical portion of population is immune to disease

Infectious agent is unable to spread in larger population due to lack of susceptible hosts

Ideal characteristics of an effective vaccine

Safe, few side effects, give long-lasting protection, ideally be low cost, stable, easy to administer (important for widespread use)

Not always achieved but what we want

Vaccines typically fit one of two categories

Attenuated or inactivated

Attenuated vaccines

Agent used can replicate (alive)

Stronger immune response … can potentially cause disease

Inactivated vaccines

Agent used cannot replicate (not alive)

Weaker immune response… cannot cause infection

Attenuated vaccines characteristics

Attenuated viruses/ bacteria

Injection, oral or nasal

No need for adjuvant

IgG response; IgA if administered orally/ nasally

Good cell-mediated immune response

Longer duration of protection

One or two doses

Very low risk of mutation to virulence

Can have significant risk to immunocompromised recipient

Poor stability in warm temperatures

Inactivated vaccines characteristics

Inactivated whole agents, toxoids, subunits, VLPs (virus-like particles), polysaccharides, conjugates

Injection

Need for adjuvant

IgG response

Poor cell-mediated immune response

Shorter protection

Multiple doses

No risk of mutation

No risk to immunocompromised recipient Poor

Good stability in warm temperatures

Important point about attenuated vaccines

Although the pathogen or agent is ‘alive’ it has been weakened to prevent strong immune response and disease

Eg induce mutations/ genetically manipulate to replace virulent genes… lowers pathogenicity

Examples of attenuated vaccines

Measles, mumps, rubella, chickenpox, yellow fever, rotavirus

In inactivated vaccines, pathogens cannot replicate but …

Antigens (epitopes) are still present

Types of inactivated vaccines

Inactivated whole agent vaccines

Toxoid vaccines

Subunit vaccines

Recombinant subunit vaccines

Virus-like particle vaccines

Inactivated whole agent vaccines

Contain entire pathogen but in a killed state

(Chemicals like formalin can kill pathogen but keep surface epitopes intact)

Eg flu, rabies, hepatitis A vaccines

Toxoid vaccines

Toxic part is destroyed but epitope remains

(Eg tetanus vaccine)

Subunit vaccines

Contain only part of pathogen structure (entire cell not present)

Eg acellular pertussis vaccine

Recombinant subunit vaccines

Same as subunit but produced by genetically engineered microbes

(Hepatitis B vaccine, yeast cells produce part of viral protein)

Virus-like particle vaccines

Capsid head of genetically engineered to be empty of RNA/DNA

(Human papillomavirus/ HPV vaccine)

Nucleic Acid-Based Vaccines

In more recent years been developed for COVID 19

mRNA vaccines or viral vector vaccines

mRNA vaccines

mRNA of antigen in lipid nanoparticles (LNPs)

Requires super low temperature storage

Viral vector vaccines

Antigen gene in non-pathogenic/ attenuated virus

Keep at room temperature

When are overall vaccines approved for usage by government

When benefits greatly outweigh risks

Why are some routine vaccines are administered in combination

Minimizes the number of injections required

(MMRV = measles, mumps, rubella, varicella vaccine)

Why do vaccines for a given pathogen typically continue to be developed over time

Renewed resistance from pathogen

To ensure greater accessibility (expense/ storage concerns)

Develop safer vaccination strategies (eg Polio vaccine)

How can we confirm the identity of pathogen

Culture the microbe from patient

Can test for the antigen itself in tissues or blood

Culture the microbe from patient

Using classical microbiology techniques: growth on selective media, metabolic tests, colony and cell morphology, etc

Often time consuming and sometimes not descriptive enough

Testing for the antigen itself in tissues or blood

Use immunological and molecular tests

Generally faster and more specific for pathogen identification

Immunological testing

Tests that use antibody-antigen interactions to diagnose disease

Simple immunological testing

Visual change that can be observed by eye

Skin test, agglutination reactions

More complex immunological testing

Happening on smaller scales and requiring specialized equipment

Immunoassays use labeled antibodies (combined with microscopy, enzyme activity, protein detection or cell sorting)

Immunoassay

Test that uses antibody-antigen interactions outside the pathogen/host (in vitro) to detect or qualify given antigens or antibodies

Steps of immunoassay

Binding of known antibodies identifies

Unknown pathogen

Binding of patient’s antibodies to know pathogen demonstrates

Current or previous infection of a given pathogen

Serum

Fluid portion of blood

The study of antibody-antigen interaction in blood is also called

Serology or ‘serological testing’

The “tier’ in these tests refer to the

Concentration of antibodies in the serum

How it used to calculate the titer of antibodies

Serial dilution

Serial dilution

Sample is added in 2- or 10- fold dilution

Antigen is added to each dilution

Titer concentration is the …

Reciprocal of last dilution giving positive result

What would be the titer of the antibody?

1000

Detection at higher dilutions means

More antibodies in sample

(More concentrated, requires more dilution)

Labeled antibodies can give detectable differences. The ‘label’ can be things like

A fluorescent dye, enzyme(s), radioactive isotope(s)

Direct assays

Known antibodies bind, ID the pathogen

Indirect assays

Known antigen binds, detects the antibodies present

Direct test

Indirect test

Colour changes from fluorescently labelled antibodies can also be used to ‘see’ …

Specific pathogen antibodies when pathogen is on microscope slide

(Can be used to detect rabies virus in brain tissue, used to confirm syphilis)

Enzyme-Linked Immunosorbent Assays (ELISAs)

Enzyme (usually peroxidase) converts colourless substrate to coloured product

Basis of many rapid test (pregnancy test)

Direct of indirect

Direct ELISA

Indirect ELISA

Immunoassay tests that use enzymes include

Group A strep tests, home pregnancy tests, HIV

What do I need to know

If you see immunoassay procedure… what is being detected?

The antigen/pathogen? (Direct)

The antibodies? (Indirect)

Will this tell me that the sample has the pathogen?

Or will it tell me that the sample has antibodies?

You can label the antibodies differently for different applications

Immunoassays combined with cell sorting

Uses lasers and electrical charges to sort cells

Look for coloured antigens in sorted cells

(Track progression of HIV by counting positive CD4 T cells

Antibodies can cross link antigens

Larger “mouthfuls” for phagocytic cells

Clumping of antigen-antibody is the basis of

Agglutination tests

Agglutination

Relies on cross linking larger cells/ smaller particles

Clumping = positive test result (antigen-antibody pair is present)

Blood typing (agglutination test)

Agglutination using beads with antibodies (certain streptococcus tests)

Skin tests

Localized immune response when antigen is introduced (injected into the skin)

Swollen area → positive result (shows antibodies for pathogen are present)

Eg tuberculin skin test to detect mycobacterium tuberculosis

Immunizations (in all their various forms)

Helps us prevent infection

Many diagnostic tests for pathogens rely on

Antibody-antigen interaction