Practical Applications of Immunology: Vaccines and Diagnostic Immunology

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Practical Applications of Immunology

Introduction

This chapter explores the development, types, and principles of vaccines, as well as immunological diagnostic techniques. These concepts are central to the prevention, detection, and management of infectious diseases in microbiology.

Vaccines

Historical Development of Vaccines

Variolation: An early smallpox prevention method involving inoculation with material from dried smallpox scabs, practiced in China during the 1400s–1700s.
Jenner's Experiment (1798): Used cowpox scab material to prevent smallpox, leading to the term vaccination (from vacca, meaning cow).
Vaccine: A suspension of organisms or fractions of organisms that induce immunity.

Principles and Effects of Vaccination

Vaccination provokes a primary immune response, resulting in antibody production and long-lived memory cells.
Subsequent exposure produces a rapid, intense secondary response.
Herd immunity: When most of the population is immune, outbreaks are sporadic due to a lack of susceptible individuals.

CDC-Recommended Vaccines to Prevent Bacterial Diseases

Disease(s)	Vaccine	Recommendation	Booster
H. influenzae type b meningitis	Polysaccharide from H. influenzae type b	Children 2–18 months	None recommended
Meningococcal ACWY meningitis	Purified polysaccharide from Neisseria meningitidis	Age 11–12 years	At 16 years and for people in an outbreak setting
Meningococcal B meningitis	Purified polysaccharide from Neisseria meningitidis	For people with substantial risk of infection, recommended for college freshmen	Every 2–3 years if risk remains
Pneumococcal pneumonia	Purified polysaccharide from 13 or 23 strains of Streptococcus pneumoniae	PVC13 for adults with certain chronic diseases; PVC23 for adults ≥65 years	None if first dose administered ≥24 months
Tetanus, diphtheria, pertussis	DTaP (children younger than 7 years) and Tdap (older children and adults for tetanus and pertussis)	DTaP (children 2, 4, 6, 15–18 months, 4–6 years); Tdap for adolescents (11–12 years) and adults	Tdap or Td every 10 years

CDC-Recommended Vaccines to Prevent Viral Diseases

Disease(s)	Vaccine	Recommendation	Booster
Chickenpox (varicella)	Attenuated virus	For infants aged 12 months	Duration of immunity not known
COVID-19	mRNA	Two dose series, any age	Recommended
Dengue	Attenuated recombinant virus	Age 9–16 years living in dengue-endemic areas	None recommended
Hepatitis A	Inactivated virus	Children age 1 year, those in/travel to endemic areas, those handling/working with infected non-human primates	Duration of protection at least 10 years
Hepatitis B	Virus antigen produced in yeast cells	For infants and children, for adults at risk (healthcare workers, people with multiple sex partners, etc.)	Duration of protection at least 7 years; need for boosters uncertain
Human Papillomavirus	Virus-like particle produced in yeast cells	All children ages 9–12	Duration at least 5 years
Influenza	Inactivated virus, attenuated virus, recombinant virus	Everyone over 6 months of age	Annual
Measles	Attenuated virus	For infants aged 15 months	Adults if exposed during outbreak
Mumps	Attenuated virus	For infants aged 15 months	Adults if exposed during outbreak
Poliomyelitis	Inactivated virus	For children; for adults, as risk to exposure warrants	Duration of immunity not known
Rabies	Inactivated virus	For field biologists in contact with wildlife in endemic areas; for veterinarians; for people exposed to rabies virus by bites	Every 2 years
Rotavirus	Attenuated virus	Oral, for infants up to 8 months	None recommended
Rubella	Attenuated virus	For infants aged 15 months; for females of childbearing age who are not pregnant	Adults if exposed during outbreak
Smallpox	Attenuated vaccinia virus	Certain military and health care workers	Duration of protection estimated at about 3 to 5 years
Zoster (shingles)	Virus antigen produced in mammalian cells	For adults over 50 years; for immunocompromised people over 19 years	None recommended

Types of Vaccines and Their Characteristics

Attenuated Vaccines

Contain weakened pathogens with reduced virulence.
Closely mimic natural infection; organisms replicate in the body, magnifying the immune response.
Confers lifelong immunity (both humoral and cellular).
Not recommended for immunocompromised patients due to risk of reversion to virulent form (e.g., oral polio vaccine).

Inactivated Vaccines

Contain whole microbes that are killed or inactivated.
Safer than attenuated vaccines but require repeated booster doses.
Induce mostly humoral immunity.

Subunit Vaccines

Use antigenic fragments to stimulate an immune response.
Recombinant vaccines: Produced by genetic modification of yeast or insects (e.g., Hepatitis B vaccine).
Toxoids: Inactivated toxins (e.g., diphtheria, tetanus).
Virus-like particle (VLP) vaccines: Resemble intact viruses but lack viral genetic material (e.g., HPV vaccine).

Polysaccharide and Conjugated Vaccines

Polysaccharide vaccines: Made from molecules in pathogen's capsule; not very immunogenic (e.g., pneumococcal vaccine).
Conjugated vaccines: Polysaccharide antigen attached to a protein carrier, making the vaccine immunogenic in infants.

DNA and mRNA Vaccines

DNA vaccines: Injected DNA directs synthesis of protein antigen, stimulating humoral and cellular immunity.
mRNA vaccines: mRNA in lipid nanoparticles directs synthesis of encoded antigen (e.g., COVID-19 vaccines).

Recombinant Vector Vaccines

Avirulent viruses or bacteria genetically modified to deliver DNA coding for antigens.

Vaccine Production, Administration, and Safety

Vaccine Production

DNA, mRNA, and recombinant vector vaccines do not require animal hosts for pathogen growth.
Plants are being explored as potential production systems.

Adjuvants

Additives that enhance vaccine effectiveness (e.g., alum, monophosphoryl lipid A).
Improve innate immune response via Toll-like receptor activation.

Vaccine Administration

Oral vaccines are favored for ease and effectiveness against GI pathogens (e.g., rotavirus, cholera).
Nasal vaccines (e.g., attenuated influenza vaccine) and skin patch vaccines (e.g., Nanopatch™) are alternative delivery methods.
Multiple-combination vaccines allow immunization against several diseases simultaneously.

Vaccine Safety

Rarely, vaccines may cause disease (e.g., variolation, oral polio vaccine).
No scientific evidence links MMR vaccines to autism.
Vaccines are the safest and most effective means of preventing infectious diseases in children.
mRNA vaccines have an outstanding safety record for COVID-19 prevention.

Diagnostic Immunology

Principles of Diagnostic Immunology

Sensitivity: Probability that a test is reactive if the specimen is a true positive.
Specificity: Probability that a positive test will not be reactive if the specimen is a true negative.
Immunologic-based diagnostic tests rely on interactions between antibodies and antigens.
Known antibodies can identify unknown pathogens; known pathogens can detect unknown antibodies.

Use of Monoclonal Antibodies

Hybridoma: Fusion of a cancerous B cell (myeloma) with a normal antibody-producing B cell, producing monoclonal antibodies (Mabs).
Mabs are uniform, highly specific, and produced in large quantities.
Applications include diagnostic tools and therapies for diseases such as multiple sclerosis, Crohn's disease, psoriasis, cancer, asthma, arthritis, and COVID-19.
Types of monoclonal antibodies:
- Chimeric: Mouse variable region, human constant region.
- Humanized: Mostly human, except for mouse antigen-binding sites.
- Fully human: Produced from mice with human antibody genes.

Precipitation Reactions

Soluble antigens react with IgG or IgM antibodies to form large, interlocking aggregates (lattices) that precipitate from solution.
Precipitin reactions occur only at optimal antigen-antibody ratios.
Precipitin ring test: Cloudy line forms at the optimal ratio.
Immunodiffusion tests: Precipitation reactions in agar gel; precipitate forms at optimal ratio.
Immunoelectrophoresis: Combines electrophoresis with immunodiffusion to separate serum proteins.

Agglutination Reactions

Particulate antigens bind to antibodies, forming visible aggregates.
Direct agglutination tests: Detect antibodies against large cellular antigens; measure antibody concentration (titer).
Seroconversion: Significant change in titer as disease progresses.
Indirect (passive) agglutination tests: Antibody reacts with soluble antigen attached to particles (e.g., latex beads).
Hemagglutination: RBC surface antigens agglutinate with antibodies; used in blood typing.

Neutralization Reactions

Neutralization: Antigen-antibody reaction blocks harmful effects of bacterial exotoxins or prevents viruses from infecting cells.
Virus neutralization test: Detects antibodies by their ability to prevent cytopathic effects in cell cultures or eggs; identifies virus and determines antibody titer.
Viral hemagglutination inhibition test: Used for subtyping viruses; antibodies in serum inhibit hemagglutination.

Complement-Fixation Reactions

Complement fixation: Complement serum protein binds to antigen-antibody complex; detects small amounts of antibody.
Useful for diagnosing viral, fungal, and rickettsial diseases.

Fluorescent-Antibody Techniques

Antibodies are combined with fluorescent dyes and visualized using a fluorescence microscope.
Direct FA tests: Identify microorganisms in clinical specimens.
Indirect FA tests: Detect specific antibodies in serum using fluorescein-labeled anti-human immune serum globulin.
Fluorescence-activated cell sorter (FACS): Labels cells with fluorescent antibodies, sorts cells by size and fluorescence; used to enumerate CD4+ T cells in AIDS progression.

Enzyme-Linked Immunosorbent Assay (ELISA)

Direct ELISA: Detects antigens by mixing sample with antibody and enzyme-linked antibodies; color change indicates presence.
Indirect ELISA: Detects antibodies in a sample.

Western Blotting (Immunoblotting)

Proteins are separated by electrophoresis, transferred to a membrane, and detected using enzyme-linked antibodies and substrate for color visualization.

Rapid Antigen Tests

SARS-CoV-2 rapid antigen test: Lateral flow assay using nasal swab; antigens captured by labeled antibody form a colored line.

Big Picture: Vaccine-Preventable Diseases

Impact of Immunization Campaigns

Vaccines have saved millions of lives (e.g., diphtheria: >15,000 deaths in 1921 vs. 14 cases since 1999).
Some diseases persist due to low vaccination rates, poverty, lack of infrastructure, need for boosters, and safety concerns.
Global initiatives aim to eliminate diseases (e.g., Measles and Rubella Initiative, Global Polio Eradication Initiative).