Chapter 17 Immunization and Immune Testing Microbiology Chapter 17 Immunization and Immune Testing

Immunization Two Artificial Methods of Immunity Active immunization Administration of antigens so patient actively mounts an adaptive immune response Passive immunotherapy Individual acquires immunity through the transfer of antibodies formed by immune individual or animal

Immunization Brief History of Immunization Chinese noticed children who recovered from smallpox did not contract the disease again. They infected children with material from a smallpox scab to induce immunity. Process known as variolation Variolation spread to England and America but was stopped due to risk of death.

Immunization Brief History of Immunization 1796—Edward Jenner discovered process of vaccination 1879—Louis Pasteur developed a vaccine against Pasteurella Antibody transfer developed when it was discovered vaccines protected through the action of antibodies

Figure 17.1 The effect of immunization in reducing the prevalence of two infectious diseases in the United States. 5

Immunization Brief History of Immunization Many developing nations do not receive vaccines Effective vaccines not developed for some pathogens Vaccine-associated risks discourage investment in developing new vaccines

Immunization Active Immunization Vaccine types Attenuated (modified live) vaccines Use pathogens with reduced virulence Process of reducing virulence called attenuation Can result in mild infections Active microbes stimulate a strong immune response Can provide contact immunity Modified microbes may retain enough residual virulence to cause disease in susceptible individuals

Immunization Active Immunization Vaccine types Inactivated (killed) vaccines Safer than live vaccines Whole agent vaccines Inactivated but whole microbes Subunit vaccines Antigenic fragments of microbes Often require multiple doses to achieve full immunity Often contain adjuvants Chemicals added to increase effective antigenicity

Immunization Active Immunization Vaccine types Toxoid vaccines Chemically or thermally modified toxins used to stimulate active immunity Useful for some bacterial diseases Stimulate antibody-mediated immunity Require multiple doses because toxoids possess few antigenic determinants

Immunization Active Immunization Vaccine types Combination vaccines Simultaneous administration of antigens from several pathogens Vaccines using recombinant gene technology Research attempts to make vaccines more effective, cheaper, and safer Recombinant DNA techniques used to improve vaccines

Immunization Active Immunization Vaccine manufacture Many vaccines mass-produced by growing microbes in culture vessels Viruses are cultured inside chicken eggs Individuals with egg allergies must avoid some vaccines

Figure 17.3 The CDC’s recommended immunization schedule for the general population. 12

Table 17.1 Principal Vaccines to Prevent Human Diseases (1 of 2) 13

Table 17.1 Principal Vaccines to Prevent Human Diseases (2 of 2) 14

Immunization Active Immunization Vaccine safety Problems associated with immunization Mild toxicity Risk of anaphylactic shock Residual virulence from attenuated viruses Allegations certain vaccines cause autism, diabetes, and asthma Research has not substantiated these allegations

Immunization Passive Immunotherapy Administration of antiserum that contains preformed antibodies Provides immediate protection against a recent infection or ongoing disease Antisera have several limitations Can trigger allergic reactions called serum sickness Antibodies of antisera are degraded relatively quickly Individual not protected from subsequent infections

Figure 17.5 The characteristics of immunity produced by active immunization (red) and passive immunotherapy (green). 17

Serological Tests That Use Antigens and Corresponding Antibodies Serology is the determination of the presence of specific antigens or antibodies in blood serum Serological tests available to identify a variety of antigens and antibodies in serum Serological tests have several uses Monitor the spread of infection within a population Establish diagnosis of disease

Serological Tests That Use Antigens and Corresponding Antibodies Precipitation Tests One of the simplest of serological tests Antigens and antibody mixed in the proper proportion form large complexes called precipitates Antigen-antibody complexes also called immune complexes Immunodiffusion is a common precipitation technique

Figure 17.6 Characteristics of precipitation reactions. 20

Figure 17.7 Immunodiffusion, a type of precipitation reaction. 21

Serological Tests That Use Antigens and Corresponding Antibodies Turbidimetric and Nephelometric Tests Turbidimetry and nephelometry measure the cloudiness of a solution Turbidimetry measures the light passing through a solution Nephelometry measures the light reflected from a solution Can be used to quantify the amounts of proteins in serum

Serological Tests That Use Antigens and Corresponding Antibodies Agglutination Tests Agglutination occurs due to the cross-linking of antibodies with particulate antigens Agglutination is the clumping of insoluble particles Precipitation involves the aggregation of soluble molecules Reactions can be easy to see and interpret with the unaided eye Hemagglutination Agglutination of red blood cells Can be used to determine blood type

Figure 17.8 The use of hemagglutination to determine blood types in humans. 24

Serological Tests That Use Antigens and Corresponding Antibodies Agglutination Tests Titration Method to measure antibody levels in blood sera Serum being tested is serially diluted and tested for agglutinating activity Highest dilution of serum giving a positive reaction is the titer

Figure 17.9 Titration, the use of agglutination to quantify the amount of antibody in a serum sample. 26

Serological Tests That Use Antigens and Corresponding Antibodies Neutralization Tests Viral neutralization Cytopathic effect Viruses introduced into appropriate cell cultures will kill the cells Ability of virus to kill culture cells is neutralized when virus is first mixed with antibodies against it Viral neutralization test Mixture of virus and serum added to cell culture Absence of cytopathic effect indicates presence of antibodies against the virus in the serum Identifies whether individual exposed to a particular virus

Serological Tests That Use Antigens and Corresponding Antibodies Neutralization Tests Viral hemagglutination inhibition test Useful for viruses that aren’t cytopathic Based on viral hemagglutination Some viral surface proteins can clump red blood cells A serum sample that contains antibodies against a specific virus will inhibit viral hemagglutination Commonly used to detect antibodies against influenza, measles, and mumps

Serological Tests That Use Antigens and Corresponding Antibodies The Complement Fixation Test Based on generation of membrane attack complexes during complement activation Used to detect the presence of specific antibodies in an individual’s serum Can detect antibody amounts too small to detect by agglutination Replaced by other serological methods

Serological Tests That Use Antigens and Corresponding Antibodies Labeled Antibody Tests Use antibody molecules linked to some “label” that enables them to be easily detected Used to detect either antigens or antibodies

Serological Tests That Use Antigens and Corresponding Antibodies Labeled Antibody Tests Fluorescent antibody tests Use fluorescent dyes as labels Fluorescently labeled antibodies used in two types of tests Direct fluorescent antibody tests Indirect fluorescent antibody tests

Figure 17.10 Here rabies proteins (green) are revealed in tissue. 32

Figure 17.11 The indirect fluorescent antibody test. 33

Serological Tests That Use Antigens and Corresponding Antibodies Labeled Antibody Tests ELISAs (EIAs) Stands for enzyme-linked immunosorbent assay (ELISA) or enzyme immunoassay (EIA) Uses an enzyme as the label Reaction of enzyme with its substrate produces a colored product indicating a positive test Commonly used to detect the presence of serum antibodies

Figure 17.12 Enzyme-linked immunosorbent assay (ELISA), also known as enzyme immunoassay (EIA). 35

Serological Tests That Use Antigens and Corresponding Antibodies Labeled Antibody Test ELISAs (EIAs) Advantages of the ELISA Can detect either antibody or antigen Sensitive Can quantify amounts of antigen or antibody Easy to perform and can test many samples quickly Relatively inexpensive and easy to automate Plates coated with antigen can be stored for later testing

Serological Tests That Use Antigens and Corresponding Antibodies Labeled Antibody Test ELISAs (EIAs) Antibody sandwich ELISA Modification of the ELISA technique Commonly used to detect antigen Antigen being tested for is “sandwiched” between two antibody molecules

Figure 17.13 An antibody sandwich ELISA. 38

Serological Tests That Use Antigens and Corresponding Antibodies Labeled Antibody Test Immunoblot Also called a western blot Technique to detect antibodies against multiple antigens Used to confirm the presence of proteins Three steps Electrophoresis separates proteins in a solution Blotting transfers protein to nitrocellulose membrane Detects color where antibody has bound to proteins

Figure 17.14 Immunoblotting (western blotting). 40

Serological Tests That Use Antigens and Corresponding Antibodies Point-of-Care Testing Simple immunoassays that give results in minutes Useful in determining a quick diagnosis Common tests Immunofiltration assay Immunochromatography assay

Serological Tests That Use Antigens and Corresponding Antibodies Point-of-Care Testing Immunofiltration Rapid ELISA that uses antibodies bound to membrane filters rather than plates Reduced time to complete the assay Due to the large surface area of the membrane filter

Serological Tests That Use Antigens and Corresponding Antibodies Recent Developments in Immune Testing Immunochromatography Very rapid and easy-to-read ELISAs Antigen solution flows through a porous strip Encounters labeled antibody Visible line produced when antigen-antibody immune complexes encounter antibody against them Used in pregnancy testing and for rapid identification of some infections

Figure 17.15 Immunochromatographic dipstick. 44

Table 17.2 Antibody-Antigen Immunological Tests and Some of Their Uses 45