Immunization pdf - D. Talib

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

476

n expanded knowledge of immune function has yielded sig-
nificant breakthroughs in medical technology for manipulat-
ing and monitoring the immune system. One of the most prac-

tical  benefits  of  immunology  is  administering  vaccines  and  other
immune treatments against common infectious diseases such as hep-
atitis and tetanus that once caused untold sickness and death. Another
valuable application of this technology is testing the blood for signs of
infection or disease. It is routine medical practice to diagnose such in-
fections as HIV, syphilis, hepatitis B, and rubella by means of immuno-
logic  analysis.  Both  separately  and  in  combination,  these  methods
have made sweeping contributions to individual and community health
by  improving  diagnosis  and  treatment,  and  controlling  the  spread  of
disease. Many hopes for the survival of humankind lie in our ability to
harness the amazing workings of the immune system.

Chapter Overview

• Discoveries in basic immune function have created powerful medical

tools to artificially induce protective immunities and to determine the

status of the immune system.

• Immunization may be administered by means of passive and active

methods.

• Passive immunity is acquired by infusing antiserum taken from other

patients’ blood that contains high levels of protective antibodies. This

form of immunotherapy is short-lived.

• Active methods involve administering a vaccine against an infectious

agent that may be encountered in the future. This form of protection

provides a longer-lived immunity.

• Vaccines contain some form of microbial antigen that has been altered

so that it stimulates a protective immune response without causing the

disease.

• Vaccines can be made from whole dead or live cells and viruses, parts

of cells or viruses, or by recombinant DNA techniques.

• Reactions between antibodies and antigens provide specific and

sensitive tests that can be used in diagnosis of disease and

identification of pathogens.

• Serology involves the testing of a patient’s blood serum for antibodies

that can indicate a current or past infection and the degree of immunity.

• Tests that produce visible interactions of antibodies and antigens

include agglutination, precipitation, and complement fixation.

• Assays can be used to separate antigens and antibodies and visualize

them with radioactivity or fluorescence. These include

immunoelectrophoresis, the Western blot, and direct and indirect

immunoassays.

Practical Applications of Immunologic
Function

A knowledge of the immune system and its responses to antigens
has provided extremely valuable biomedical applications in two
major areas: (1) use of antiserum and vaccination to provide artifi-
cial protection against disease and (2) diagnosis of disease through
immunologic testing.

Detail from “The Cowpock,” an 1808 etching that caricatured the worst fears
of the English public concerning Edward Jenner’s smallpox vaccine.

Immunization and
Immune Assays

CHAPTER

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

Practical Applications of Immunologic Function

477

IMMUNIZATION: METHODS OF MANIPULATING
IMMUNITY FOR THERAPEUTIC PURPOSES

The concept of artificially induced immunity was introduced in
chapters 14 and 15. Methods that actively or passively immunize
people are widely used in disease prevention and treatment. In the
case of passive immunization, a patient is given preformed anti-
bodies, which is actually a form of

immunotherapy. In the case of

active immunization, a patient is vaccinated with a microbe or its
antigens, providing a form of advance protection.

Immunotherapy: Artificial Passive Immunity

The first attempts at passive immunization involved the transfusion
of horse serum containing antitoxins to prevent tetanus and to treat
patients exposed to diphtheria. Since then, antisera from animals
have been replaced with products of human origin that function
with various degrees of specificity.

Immune serum globulin (ISG),

sometimes called

gamma globulin, contains immunoglobulin ex-

tracted from the pooled blood of at least 1,000 human donors. The
method of processing ISG concentrates the antibodies to increase
potency and eliminates potential pathogens (such as the hepatitis B
and HIV viruses). It is a treatment of choice in preventing measles
and hepatitis A and in replacing antibodies in immunodeficient pa-
tients. Most forms of ISG are injected intramuscularly to minimize
adverse reactions, and the protection it provides lasts 2 to 3 months.

A preparation called

specific immune globulin (SIG) is de-

rived from a more defined group of donors. Companies that prepare
SIG obtain serum from patients who are convalescing and in a hy-
perimmune state after such infections as pertussis, rabies, tetanus,
chickenpox, and hepatitis B. These globulins are preferable to ISG
because they contain higher titers of specific antibodies obtained
from a smaller pool of patients. Although useful for prophylaxis in
persons who have been exposed or may be exposed to infectious
agents, these sera are often limited in availability.

When a human immune globulin is not available, antisera

and antitoxins of animal origin can be used. Sera produced in
horses are available for diphtheria, botulism, and spider and snake
bites. Unfortunately, the presence of horse antigens can stimulate
allergies such as serum sickness or anaphylaxis (see chapter 17).
Although donated immunities only last a relatively short time, they
act immediately and can protect patients for whom no other useful
medication or vaccine exists.

Artificial Active Immunity: Vaccination

Active immunity can be conferred artificially by

vaccination—

exposing a person to material that is antigenic but not pathogenic.
The discovery of vaccination was one of the farthest reaching and
most important developments in medical science (Historical High-
lights 16.1). The basic principle behind vaccination is to stimulate

HISTORICAL HIGHLIGHTS

16.1

The Lively History of Active Immunization

The basic notion of immunization has existed for thousands of years. It
probably stemmed from the observation that persons who had recovered
from certain communicable diseases rarely if ever got a second case. Un-
doubtedly, the earliest crude attempts involved bringing a susceptible
person into contact with a diseased person or animal. The first recorded
attempt at immunization occurred in sixth century China. It consisted of
drying and grinding up smallpox scabs and blowing them with a straw
into the nostrils of vulnerable family members. By the tenth century, this
practice had changed to the deliberate inoculation of dried pus from the
smallpox pustules of one patient into the arm of a healthy person, a tech-
nique later called

variolation (variola is the smallpox virus). This

method was used in parts of the Far East for centuries before Lady Mary
Montagu brought it to England in 1721. Although the principles of the
technique had some merit, unfortunately, many recipients and their con-
tacts died of smallpox. This outcome vividly demonstrates a cardinal rule
for a workable vaccine: It must contain an antigen that will provide pro-
tection but not cause the disease. Variolation was so controversial that
any English practitioner caught doing it was charged with a felony.

Eventually, this human experimentation paved the way for the first

really  effective  vaccine,  developed  by  the  English  physician  Edward
Jenner in 1796 (see chapter 24). Jenner conducted the first scientifically
controlled  study,  one  that  had  a  tremendous  impact  on  the  advance  of
medicine.  His  work  gave  rise  to  the  words

vaccine and vaccination

(from L.,

vacca, cow), which now apply to any immunity obtained by in-

oculation with selected antigens. Jenner was inspired by the case of a

dairymaid who had been infected by a pustular infection called cowpox.
This is a related virus that afflicts cattle but causes a milder condition in
humans. She explained that she and other milkmaids had remained free
of smallpox. Other residents of the region expressed a similar confidence
in the cross-protection of cowpox. To test the effectiveness of this new
vaccine, Jenner prepared material from human cowpox lesions and inoc-
ulated a young boy. When challenged 2 months later with an injection of
crusts from a smallpox patient, the boy proved immune.

Jenner’s discovery—that a less pathogenic agent could confer

protection against a more pathogenic one—is especially remarkable in
view of the fact that microscopy was still in its infancy and the nature of
viruses was unknown. At first, the use of the vaccine was regarded with
some fear and skepticism (see chapter opening illustration). When Jen-
ner’s method proved successful and word of its significance spread, it
was eventually adopted in many other countries. Eventually, the original
virus mutated into a unique strain (

vaccina virus) that was the modern ba-

sis of the vaccine. Now that smallpox is no longer a threat, smallpox vac-
cination has been essentially discontinued in most areas.

Other historical developments in vaccination included using heat-

killed bacteria in vaccines for typhoid fever, cholera, and plague and
techniques for using neutralized toxins for diphtheria and tetanus.
Throughout the history of vaccination, there have been vocal opponents
and minimizers, but numbers do not lie. Whenever a vaccine has been in-
troduced, the prevalence of that disease has declined.

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

478

CHAPTER 16 Immunization and Immune Assays

a  primary  and  secondary  anamnestic  response  (see  figure  15.18)
that  primes  the  immune  system  for  future  exposure  to  a  virulent
pathogen.  If  this  pathogen  enters  the  body,  the  immune  response
will be immediate, powerful, and sustained.

Vaccines have profoundly reduced the prevalence and impact

of many infectious diseases that were once common and often
deadly. In this section, we survey the principles of vaccine prepara-
tion and important considerations surrounding vaccine indication
and safety. (Vaccines are also given specific consideration in later
chapters on bacterial and viral diseases.)

Principles of Vaccine Preparation

A vaccine must be con-

sidered from the standpoints of antigen selection, effectiveness,
ease in administration, safety, and cost. In natural immunity, an
infectious agent stimulates appropriate B and T lymphocytes and
creates memory clones. In artificial active immunity, the objec-
tive is to obtain this same response with a modified version of the
microbe or its components. A safe and effective vaccine should
mimic the natural protective response, not cause a serious infec-
tion  or  other  disease,  have  long-lasting  effects  in  a  few  doses,
and be easy to administer. Most vaccine preparations contain one
of  the  following  antigenic  stimulants  (figure  16.1):  (1)  killed
whole  cells  or  inactivated  viruses,  (2)  live,  attenuated  cells  or
viruses, (3) antigenic components of cells or viruses, or (4) ge-
netically engineered microbes or microbial antigens. A survey of
the major licensed vaccines and their indications is presented in
table 16.1.

Large, complex antigens such as whole cells or viruses are

very effective immunogens. Depending on the vaccine, these are
either killed or attenuated.

Killed or inactivated vaccines are

prepared by cultivating the desired strain or strains of a bac-
terium or virus and treating them with formalin, radiation, heat,
or some other agent that does not destroy antigenicity. One type
of vaccine for the bacterial disease typhoid fever is of this type
(see chapter 20). Salk polio vaccine and influenza vaccine con-
tain inactivated viruses. Because the microbe does not multiply,
killed vaccines often require a larger dose and more boosters to
be effective.

A number of vaccines are prepared from

live, attenuated*

microbes.

Attenuation is any process that substantially lessens or

negates the virulence of viruses or bacteria. It is usually achieved
by  modifying  the  growth  conditions  or  manipulating  microbial
genes in a way that eliminates virulence factors. Attenuation meth-
ods include long-term cultivation, selection of mutant strains that
grow at colder temperatures (cold mutants), passage of the microbe
through unnatural hosts or tissue culture, and removal of virulence
genes. The vaccine for tuberculosis (BCG) was obtained after 13
years of subculturing the agent of bovine tuberculosis (see chapter
19). Vaccines for measles, mumps, polio (Sabin), and rubella con-
tain live, nonvirulent viruses. The advantages that favor live prepa-
rations  are:  (1) Viable  microorganisms  can  multiply  and  produce
infection (but not disease) like the natural organism; (2) they confer
long-lasting  protection;  and  (3)  they  usually  require  fewer  doses
and boosters than other types of vaccines. Disadvantages of using

live microbes in vaccines are that they require special storage facil-
ities, can be transmitted to other people, and can mutate back to a
virulent strain (see polio, chapter 25).

If the exact antigenic determinants that stimulate immunity

are known, it is possible to produce a vaccine based on a selected
component of a microorganism. These vaccines for bacteria are
called

acellular or subcellular vaccines. For viruses, they are

(a) Whole-Cell Vaccines

Killed cell or
inactivated
virus

Dead, but
antigenicity
is retained

Heat or

chemicals

Vaccine
stimulates
immunity
but pathogen
cannot multiply.

Administer

Live, attenuated
cells or viruses

Alive, with
same antigenicity

Virulence

is eliminated

or reduced

Administer

Vaccine
can multiply and
boost immune
stimulation.

Ags

(b) Acellular or Subunit Vaccine

Hepatitis B virus

Surface Ag

Plasmid with gene
that codes for
surface Ag

Yeast cloning vector

Synthesis and release
of surface antigen

FIGURE 16.1

Strategies in vaccine design.

(a) Whole cells or viruses, killed

or attenuated. (b) Acellular or subunit vaccines are made by

disrupting  the  microbe  to  release  various  molecules  or  cell  parts  that
can  be  isolated  and  purified.  (c) Recombinant  vaccines  are  made  by
isolating  a  gene  for  antigenicity  from  the  pathogen  (here  a  hepatitis
virus)  and  splicing  it  into  a  plasmid.  Insertion  of  the  recombinant
plasmid  into  a  cloning  host  (yeast)  results  in  the  production  of  large
amounts  of  viral  surface  antigen  to  use  in  vaccine  preparation.

attenuated

(ah-ten´-yoo-ayt-ed) L.

attenuare, to thin. Able to multiply, but nonvirulent.

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

Practical Applications of Immunologic Function

479

called

subunit vaccines. The antigen used in these vaccines may be

taken from cultures of the microbes, produced by rDNA technol-
ogy, or synthesized chemically.

Examples of component antigens currently in use are the

capsules of the pneumococcus and meningococcus, the protein
surface antigen of anthrax, and the surface proteins of hepatitis B
virus. A special type of vaccine is the

toxoid,* which consists of a

purified bacterial exotoxin that has been chemically denatured. By

eliciting the production of antitoxins that can neutralize the natural
toxin, toxoid vaccines provide protection against toxinoses such as
diphtheria and tetanus.

New Vaccine Strategies

Despite  considerable  successes,  dozens  of  bacterial,  viral,  proto-
zoan, and fungal diseases still remain without a functional vaccine.
Of  all  of  the  challenges  facing  vaccine  specialists,  probably  the
most difficult has been choosing a vaccine antigen that is safe and
that  properly  stimulates  immunity.  Currently,  much  attention  is

TABLE 16.1

Currently Approved Vaccines

Disease/Preparation

Route of Administration

Recommended Usage/Comments

Contain Killed Whole Bacteria

Cholera

Subcutaneous (SQ) injection

For travelers; effect not long-term

Typhoid

Intramuscular (IM)

For travelers only; efficacy variable

Plague

For exposed individuals and animal workers; variable protection

Contain Live, Attenuated Bacteria

Tuberculosis (BCG)

Intradermal (ID) injection

For high-risk occupations only; protection variable

Acellular Vaccines (Capsular Polysaccharides)

Meningitis (meningococcal)

For protection in high-risk infants, military recruits; short duration

Meningitis (

Haemophilus influenzae) IM

For infants and children; may be administered with DTaP

Pneumococcal pneumonia

IM or SQ

Important for people at high risk: the young, elderly, and

immunocompromised; moderate protection

Pertussis (aP)

For newborns and children; contains recombinant protein antigens

Toxoids (Formaldehyde-Inactivated Bacterial Exotoxins)

Diphtheria

A routine childhood vaccination; highly effective in systemic protection

Tetanus

A routine childhood vaccination; highly effective

Botulism

Only for exposed individuals such as laboratory personnel

Contain Inactivated Whole Viruses

Poliomyelitis (Salk)

Routine childhood vaccine; now used as first choice

Rabies

For victims of animal bites or otherwise exposed; effective

Influenza

For high-risk populations; requires constant updating for new strains;

immunity not durable

Hepatitis A

Protection for travelers, institutionalized people

Contain Live, Attenuated Viruses

Adenovirus infection

Oral

For immunizing military recruits

Measles (rubeola)

Routine childhood vaccine; very effective

Mumps (parotitis)

Routine childhood vaccine; very effective

Poliomyelitis

Oral

Routine childhood vaccine; very effective, but can cause polio

Rubella

Routine childhood vaccine; very effective

Chickenpox (varicella)

Routine childhood vaccine; immunity can diminish over time

Yellow fever

Travelers, military personnel in endemic areas

Subunit Viral Vaccines

Hepatitis B

Recommended for all children, starting at birth; also for health workers

and others at risk

Influenza

See influenza above

Recombinant Vaccines

Hepatitis B

Used more often than subunit, but for same groups

Pertussis

See acellular above

Lyme Disease (LymeRIX)

Made from the surface protein; used for persons with increased risk of

exposure to ticks

toxoid

(tawks´-oyd) Toxinlike.

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

480

CHAPTER 16 Immunization and Immune Assays

being focused on newer strategies for vaccine preparation that
employ antigen synthesis, recombinant DNA, and gene cloning
technology.

When the exact composition of an antigenic determinant is

known, it is possible to synthesize it. This ability permits preserva-
tion of antigenicity while greatly increasing antigen purity and con-
centration. The malaria vaccine currently being used in areas of
South America and Africa is composed of three synthetic peptides
from the parasite. Several biotechnology companies are exploring
the possibility of using plants to synthesize microbial proteins, po-
tentially leading to mass production of edible vaccine antigens.

Some of the genetic engineering concepts introduced in

chapter 10 offer novel approaches to vaccine development. These
methods are particularly effective in designing vaccines for obli-
gate parasites that are difficult or expensive to culture, such as the
syphilis spirochete or the malaria parasite. This technology pro-
vides a means of isolating the genes that encode various microbial
antigens, inserting them into plasmid vectors, and cloning them in
appropriate hosts. The outcome of recombination can be varied as
desired. For instance, the cloning host can be stimulated to synthe-
size and secrete a protein product (antigen), which is then harvested
and purified (figure 16.1

c). This is how certain vaccines for hepati-

tis B rotavirus and Lyme disease are prepared. Antigens from the
agents of syphilis,

Schistosoma, and influenza have been similarly

isolated and cloned and are currently being considered as potential
vaccine material.

Another ingenious technique using genetic recombination

has been nicknamed the

Trojan horse vaccine. The term derives

from an ancient legend in which the Greeks sneaked soldiers into
the fortress of their Trojan enemies by hiding them inside a large,
mobile wooden horse. In the microbial equivalent, genetic material
from  a  selected  infectious  agent  is  inserted  into  a  live  carrier
microbe that is nonpathogenic. In theory, the recombinant microbe
will  multiply  and  express  the  foreign  genes,  and  the  vaccine
recipient  will  be  immunized  against  the  microbial  antigens.
Vaccinia, the virus originally used to vaccinate for smallpox, and
adenoviruses  have  proved  practical  agents  for  this  technique.
Vaccinia is used as the carrier in one of the experimental vaccines
for AIDS (see figure 25.19), herpes simplex 2, leprosy, and tuber-
culosis.

DNA vaccines are being hailed as the most promising of all

of the newer approaches to immunization. The technique in these
formulations is very similar to gene therapy as described in figure
10.13, except in this case, microbial (not human) DNA is inserted
into a plasmid vector and inoculated into a recipient (figure 16.2

a).

The expectation is that the human cells will take up some of the
plasmids and express the microbial DNA in the form of proteins.
Because these proteins are foreign, they will be recognized during
immune surveillance and cause B and T cells to be sensitized and
form memory cells.

Experiments with animals have shown that these vaccines

are very safe and that only a small amount of the foreign antigen
need be expressed to produce effective immunity. Another advan-
tage to this method is that any number of potential microbial pro-
teins can be expressed, making the antigenic stimulus more com-
plex and improving the likelihood that it will stimulate both
antibody and cell-mediated immunity. At the present time, over

30 DNA-based vaccines are being tested in animals. Vaccines for
Lyme disease, hepatitis C, herpes simplex, influenza, tuberculosis,
papillomavirus, and malaria are undergoing animal trials, most
with encouraging results.

Vaccine effectiveness relies, in part, on the production of

antibodies that closely fit the natural antigen. Realizing that such
reactions are very much like a molecular jigsaw puzzle, researchers
have proposed an entirely new concept in vaccines. The

anti-

idiotype vaccine is based on the principle that the antigen binding
(variable) region, or

idiotype,* of a given antibody (A) can be

antigenic  to  a  genetically  different  recipient  and  can  cause  that
recipient’s  immune  system  to  produce  antibodies  (B;  also  called
anti-idiotypic  antibodies)  specific  for  the  variable  region  on  anti-
body  A (figure  16.2

b). The purpose for making anti-idiotypic

antibodies is that they will display an identical configuration as the
desired antigen and can be used in vaccines. This method avoids
administering a microbial antigen, thus reducing the potential for
dangerous side effects. Using monoclonal antibodies, this approach
has been used to mimic the surface antigen of hepatitis B virus and
Trypanosoma with some success.

Route of Administration and Side Effects
of Vaccines

Most vaccines are injected by subcutaneous, intramuscular, or intra-
dermal routes. Oral vaccines are available for only two diseases
(table 16.1), but they have some distinct advantages. An oral dose
of a vaccine can stimulate protection (IgA) on the mucous mem-
brane of the portal of entry. Oral vaccines are also easier to give,
more readily accepted, and well tolerated. Some vaccines require
the addition of a special binding substance, or

adjuvant.* An

adjuvant  is  any  compound  that  enhances  immunogenicity  and
prolongs antigen retention at the injection site. The adjuvant pre-
cipitates  the  antigen  and  holds  it  in  the  tissues  so  that  it  will  be
released gradually. Its gradual release, presumably, facilitates con-
tact  with  macrophages  and  lymphocytes.  Common  adjuvants  are
alum (aluminum hydroxide salts), Freund’s adjuvant (emulsion of
mineral oil, water, and extracts of mycobacteria), and beeswax.

Vaccines must go through many years of trials in experimen-

tal  animals  and  humans  before  they  are  licensed  for  general  use.
Even after they have been approved, like all therapeutic products,
they are not without complications. The most common of these are
local  reactions  at  the  injection  site,  fever,  allergies,  and  other
adverse  reactions.  Relatively  rare  reactions  (about  1  case  out  of
220,000 vaccinations) are panencephalitis (from measles vaccine),
back-mutation to a virulent strain (from polio vaccine), disease due
to contamination with dangerous viruses or chemicals, and neuro-
logical effects of unknown cause (from pertussis and swine flu vac-
cines). Some patients experience allergic reactions to the medium
(eggs or tissue culture) rather than to vaccine antigens. Some recent
studies  have  attempted  to  link  childhood  vaccinations  to  later
development  of  diabetes,  asthma,  and  autism.  After  thorough
examination of records, epidemiologists have found no convincing
evidence for a connection to these diseases.

idiotype

(id´-ee-oh-type) Gr.

idios, own, peculiar. Another term for the antigen binding

site.

adjuvant

(ad´-joo-vunt) L.

adjuvare, to help.

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

Practical Applications of Immunologic Function

481

When known or suspected adverse effects have been de-

tected,  vaccines  are  altered  or  withdrawn.  Most  recently,  the
whole-cell pertussis vaccine was replaced by the acellular capsule
(aP)  form  when  it  was  associated  with  adverse  neurological  ef-
fects. The  live  oral  rotavirus  vaccine  had  to  be  withdrawn  when
children  experienced  intestinal  blockage.  Polio  vaccine  was
switched  from  live,  oral  to  inactivated  when  too  many  cases  of
paralytic disease occurred from back-mutated vaccine stocks. Vac-
cine  companies  are  also  phasing  out  certain  preservatives
(thimerosal) that are thought to cause allergies and other potential
side effects.

Professionals involved in giving vaccinations must under-

stand their inherent risks but also realize that the risks from the
infectious disease almost always outweigh the chance of an adverse
vaccine reaction. The greatest caution must be exercised in giving
live vaccines to immunocompromised or pregnant patients, the lat-
ter because of possible risk to the fetus.

To Vaccinate: Why, Whom, and When?

Vaccination confers long-lasting, sometimes lifetime, protection
in the individual, but an equally important effect is to protect the
public health. Vaccination is an effective method of establishing

Protein
from
pathogen

Protein expresed
on surface of cell

Nucleus

Foreign protein
of pathogen is
inserted into
cell membrane,
where it will
stimulate immune
response.

Cells of subject
accept plasmid
with pathogen's
DNA. DNA is
transcribed and
translated into
various proteins.

DNA vaccine
injected into
subject.

Rabbit Ab

(b) Technology for anti-idiotypic vaccines

Mouse B-cell clone
recognizes foreign
idiotype.

Idiotype
of Ab

Rabbit Ab (antibody A)

Anti-idiotypic Ab
(antibody B)
produced.

New antibodies with
same specificity for Ag
as original Ab A.

Injected into
human as
vaccine Ag

Ab alone
injected
into mouse

Anti-idiotype has
same configuration
as Ag in (1)

Genomic DNA
inserted into
plasmid vector;
plasmid is
amplified and
prepared as
vaccine.

DNA that codes
for protein antigen
extracted from
pathogen genome.

DNA of
pathogen

Plasmid

Plasmid
with DNA

(2)

(1)

(3)

(4)

(3)

(2)

(1)

(4)

(5)

(a) Technology for DNA vaccines

FIGURE 16.2

Other technologies useful in preparing vaccines.

(a) DNA vaccines contain all or part of the pathogen’s DNA, which is used to “infect” recipient’s

cells. Processing of the DNA leads to production of an antigen protein that can stimulate a specific response against that pathogen. (b) Anti-idiotypic
vaccines use antibodies as foreign proteins to mimic the structure of the natural antigen. See text for details.

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

482

CHAPTER 16 Immunization and Immune Assays

herd immunity in the population. According to this concept, in-
dividuals immune to a communicable infectious disease will not
be carriers, and therefore the occurrence of that microbe will be
reduced. With a larger number of immune individuals in a popu-
lation (herd), it will be less likely that an unimmunized member
of  the  population  will  encounter  the  agent.  In  effect,  collective
immunity  through  mass  immunization  confers  indirect  protec-
tion on the nonimmune (such as children). Herd immunity main-
tained  through  immunization  is  an  important  force  in  averting
epidemics.

Vaccination is recommended for all typical childhood dis-

eases for which a vaccine is available and for people in certain

special circumstances (health workers, travelers, military person-
nel). Table 16.2 outlines a general schedule for childhood immu-
nization and the indication for special vaccines. Not only are some
vaccines mixtures of antigens, but in certain cases several vaccines
are also administered simultaneously. Examples are military recr-
uits who receive as many as 15 injections within a few minutes and
children who receive boosters for DTaP and polio at the same time
they receive the MMR vaccine. Experts doubt that immune inter-
ference (inhibition of one immune response by another) is a signif-
icant problem in these instances, and the mixed vaccines are care-
fully balanced to prevent this eventuality. The main problem with
simultaneous administration is that side effects can be amplified.

TABLE 16.2

Recommended Regimen and Indications for Routine Vaccinations

Routine Schedules for Children and Adults

Vaccine

Birth

Months

4–6

Years

11–12

Years

Adults Comments

Mixed vaccines

Td is tetanus/diphtheria; T is tetanus
alone; either one should be given as
booster every 10 years.

First dose varies with disease incidence;
booster given at either 4– 6 or 11–12 years.

Schedule depends upon source of
vaccine; given with DTaP as TriHIBit

Single vaccines

Similar schedule to DTaP; injected
vaccine

Option depends upon condition of
infant; 3 doses given

Cannot be given to children <1 year old

Diphtheria,
Tetanus,
Pertussis

(DTaP)

DTaP

one dose

Td or T

Measles,

Mumps,

Rubella (MMR)

MMR

Haemophilus
influenzae
type b (Hib)

Hib

IPV

Hepatitis B (HB)

HB option 1

HB option 2

HB option 3

Chickenpox
(CPV)

CPV one dose

CPV

two

doses

Shaded bars indicate that a dose can be
given once during this time frame

Measles vaccine (Attenuvax) can be given alone

to children during epidemics or to adults immunized
before 1970.

Used in Cases of Specific Risk Due to Occupational or Other Exposure

Group Targeted

Health care personnel; people exposed through life-style

Children 2–14 years who live in areas of high prevalence

Elderly patients, children with sickle-cell anemia

Hospital, laboratory, health care workers

People whose jobs involve contact with animals (veterinarians, forest rangers); known or
suspected exposure to rabid animal; living in areas of high incidence

Travelers to endemic regions, including military recruits (varies with geographic destination)

Vaccine

Hepatitis B (Recombivax)

Hepatitis A (Havrix)

Pneumococcus (Pneumovax)

Influenza, polio, tuberculosis (BCG)

Rabies, plague, Lyme disease

Cholera, hepatitis B, hepatitis A, measles, yellow fever,
meningococcal meningitis, polio, rabies, typhoid, plague

Pneumococcus
Vaccine (PV)

DTaP

Used to protect children against otitis
media

Poliovirus (IPV)

IPV

one dose

IPV

DTaP. The diphtheria–tetanus–acellular pertussis

vaccine has replaced the DTP.

IPV—inactivated polio vaccine—is now indicated

as a safer alternative to the oral polio vaccine.

Talaro−Talaro: Foundations

in Microbiology, Fourth

Edition

16. Immunization and

Immune Assays

Text

Companies, 2002

Practical Applications of Immunologic Function

483

SEROLOGICAL AND IMMUNE TESTS: MEASURING
THE IMMUNE RESPONSE IN VITRO

The antibodies formed during an immune reaction are important in
combating infection, but they hold additional practical value. Char-
acteristics of antibodies (such as their quantity or specificity) can
reveal the history of a patient’s contact with microorganisms or
other antigens. This is the underlying basis of

serological testing.

Serology is the branch of immunology that traditionally deals with
in vitro diagnostic testing of serum. Serological testing is based on
the  familiar  concept  that  antibodies  have  extreme  specificity  for
antigens, so when a particular antigen is exposed to its specific an-
tibody, it will fit like a hand in a glove. The ability to visualize this
interaction by some means provides a powerful tool for detecting,
identifying,  and  quantifying  antibodies—or  for  that  matter,  anti-
gens.  The  scheme  works  both  ways,  depending  on  the  situation.
One  can  detect  or  identify  an  unknown  antibody  using  a  known
antigen, or one can use an antibody of known specificity to help de-
tect or identify an unknown antigen (figure 16.3). Modern serolog-
ical testing has grown into a field that tests more than just serum.
Urine,  cerebrospinal  fluid,  whole  tissues,  and  saliva  can  also  be
used  to  determine  the  immunologic  status  of  patients.  These  and
other immune tests are helpful in confirming a suspected diagnosis
or in screening a certain population for disease.

General Features of Immune Testing

The strategies of immunologic tests are diverse, and they underline
some of the brilliant and imaginative ways that antibodies and anti-
gens can be used as tools. We will summarize them under the head-
ings of agglutination, precipitation, immunodiffusion, complement
fixation, fluorescent antibody tests, and immunoassay tests. First
we will overview the general characteristics of immune testing, and
we will then look at each type separately.

The most effective serological tests have a high degree of

specificity and sensitivity (figure 16.4).

Specificity is the property

of a test to focus upon only a certain antibody or antigen and not to

Knowledge of the specific immune response has two practical

applications: (1) commercial production of antisera and vaccines and
(2) development of rapid, sensitive methods of disease diagnosis.

Artificial passive immunity usually involves administration of antiserum,

and  occasionally  B  and T  cells. Antibodies  collected  from  donors
(human  or  otherwise)  are  injected  into  people  who  need  protection
immediately. Examples  include  ISG  (immune  serum  globulin)  and  SIG
(specific  immune  globulin).

Artificial active agents are vaccines that provoke a protective immune

response  in  the  recipient  but  do  not  cause  the  actual  disease.
Vaccination  is  the  process  of  challenging  the  immune  system  with  a
specially  selected  antigen. Examples  are  (1)  killed  or  inactivated
microbes,  (2)  live,  attenuated  microbes,  (3)  subunits  of  microbes,  and
(4)  genetically  engineered  microbes  or  microbial  parts.

Vaccination programs seek to protect the individual directly through

raising the antibody titer and indirectly through the development of
herd immunity.

CHAPTER CHECKPOINTS

Patient's serum
antibody content unknown

Macroscopic
reaction

Prepared
known

antigen

Isolated colony,
identity unknown

ANTISERUM

Macroscopic
reaction

Antibodies of
known

identity

(a)

(b)

FIGURE 16.3

Basic principles of testing using antibodies and antigens.

(a) In

serological  diagnosis  of  disease,  a  blood  sample  is  scanned  for  the
presence  of  antibody  using  an  antigen  of  known  specificity.  A  positive
reaction  is  usually  evident  as  some  visible  sign,  such  as  color  change
or  clumping,  that  indicates  a  specific  interaction  between  antibody  and
antigen.  (The  reaction  at  the  molecular  level  is  rarely  observed.) (b) An
unknown  microbe  is  mixed  with  serum  containing  antibodies  of  known
specificity,  a  procedure  known  as  serotyping.  Microscopically  or
macroscopically  observable  reactions  indicate  a  correct  match  between
antibody  and  antigen  and  permit  identification  of  the  microbe.