Practical Applications of Immunology

1
Practical Applications of Immunology

• Vaccines
• Ways to detect antibodies
• Look at antigen/antibody binding

2
Objectives

• What are monoclonal antibodies and how are they
  made?
• What does titer mean?
• Know how the following tests are performed and
  how antigen and antibody are detected
• Precipitation reactions
• Agglutination reactions direct or indirect
• Hemagglutination
• Viral hemagglutination inhibition
• Complement fixation
• Fluorescent antibody techniques direct or
  indirect
• ELISA Direct and indirect

3
Principles of Diagnostic Immunology

• Physician collects a sample
• Antigen sample
• A bodily fluid that contains the infecting
  microbe or the microbes toxin
• Urine, feces, blood, skin, pus, throat swab,
  mucous, etc.
• Blood antiserum sample
• Blood antiserum contains the antibodies that the
  patient made against an infection if the patient
  is infected with the suspected pathogen then
  his/her serum has those antibodies in it.

4
Titer

• The amount of antibody in the antiserum
• Can be used to determine how far a disease has
  progressed

5
Monoclonal antibodies fuse B cells and cancer
cells
6
Diagnostic Immunology

• Many different tests require the use of
  antibodies and look at antigen and antibody
  binding
• Precipitation reactions
• Agglutination reactions direct or indirect
• Hemagglutination
• Viral hemagglutination inhibition
• Complement fixation
• Fluorescent antibody techniques direct or
  indirect
• ELISA Direct and indirect

7
Precipitation reactions (immunodiffusion)
8
Precipitation reactions band or line formed at
the zone of equivalence
Figure 18.4 - Overview
9
Agglutination Tests

• Agglutination occurs when antibody binds to more
  than one antigen at a time.

10
Agglutination tests

• Direct
• Used to detect antibodies in antiserum that are
  specific for large cellular antigens
• Antigen concentrations on the same but vary the
  amount of antibody
• Indirect
• Used to detect soluble antigen or antibodies
• Use coated latex beads

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Direct Agglutination Tests
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Agglutination
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Indirect Agglutination Tests
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Hemagglutination

• Agglutination of RBCs
• Blood is the antigen in the agglutination test
• Test used to determine blood type.
• If blood has type A antigens then it will
  agglutinate in the presence of type A antibodies.

15
Viral Hemagglutination

• Some viruses agglutinate RBCs
• Mumps, measles, influenza
• Viral hemagglutination inhibition test
• Antibodies bind to virus first inhibiting
  clumping of RBCs when blood is added.
• Test
• Take serum sample from patient and mix with
  suspected viral pathogen
• Add blood

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Viral hemagglutination inhibition
testantibodies inhibit hemagglutination
17
Complement fixation test indicate presence of
antigen/antibody binding
18
Fluorescent antibody tests

• Fluorescence is used as a label to detect if
  antigen and antibodies are bound together
• Direct fluorescence means that the antibody is
  directly labeled with a fluorescent dye and there
  is only one antibody
• Indirect fluorescence means that there are two
  antibodies and the second antibody has a
  fluorescent label

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Direct and indirect fluorescent tests
Figure 18.11 Overview
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ELISA

• Enzyme Linked Immunosorbent Assay
• Enzyme reacts with substrate to produce colored
  product
• Direct
• Enzyme is linked to an antibody (made by
  pharmaceutical company)
• Detects antigen from patient sample
• Ex. Home pregnancy test to test for hCG hormone
• hCG hormone is a protein present in urine when a
  woman is pregnant
• Indirect
• Detects antibodies

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Indirect
Direct
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Home pregnancy test