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5/9/10
IMMUNOLOGY LAB
5/9/10
Host Defenses
• 1st defense. Structures, chemicals and processes that work together to prevent pathogens from entering the body
– Skin, mucous membranes, microflora, body secretions
• 2nd defense. Cells, chemicals and processes that work together to destroy pathogens that managed to enter the body
– Phagocytosis, antimicrobial chemicals, inflammation and fever
• 3rd defense. Immune response against pathogens (specific Immunity)
– Macrophages and lymphocytes
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Immune System
• Lymphoid tissue and Associated Organs:
• Bone marrow• Produces B-cells
• Lymph nodes• Filters lymph
• Spleen• Filters blood
• Thymus• Produces T-cells
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Lymphoid System
• System of mononuclear phagocytes
• Lymphocytes• Macrophages• Dendritic cells • Plasma cells
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Immunity• Adaptive Immunity
– Naturally acquired (contact with pathogen)
– Artificially acquired (vaccination)
• Innate Immunity– You are born with
• Passive Immunity– Transfer of
antibodies– Activated T cells
from immune host • Active Immunity
– Antigen –
•
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– Second level• Third level
– Fourth level» Fifth level
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Innate Vs. Acquired
• Innate– Non-specific– Responds the same way – Includes barriers (skin and mucosa), chemicals
(histamine), and non-specific macrophages
• Acquired– Memory and specificity– Includes B-cell and T-cell mediated response– Slower response than innate
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Immune Response
Antigens Surface molecules of
pathogens Nonliving toxins,
chemicals, drugs, and foreign particles
Antibody Primary—produced in
response to antigen (ELISA) Secondary—
bind to primary antibodies
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B-cells Make Antibodies
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Production of Antibodies
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ELISA – Enzyme linked ImmunoSorbent Assay
Qualitative or quantitative
Detects antigens or antibodies
Amount of colored product from secondary antigen (HRP conjugate) is proportional to antigen-antibody interaction
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Simulating a Spread of An Infectious Disease, and Tracking Down this Source
• Shared fluids will be assayed using ELISA to
determine if you have been infected and to track
down the source of the infection!
Unknown Fluid Sample (A)
Sample A + B (mix)→ ½ AB Sample AB + C (mix) → ½ ABC Sample ABC + D (mix) → ABCD
*BE CAREFUL OF WHICH SAMPLES YOU CROSS CONTAMINATE*
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AB
1stD
3rd
**
I
HF
J
EG
A*
D*
C*
*
*
*
*
*
C*
2nd
source • In this example, A shares fluids with individual B, then C, then D• * = HIV infection• You will share fluids with three different people from the
neighboring group
Infection Simulation
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Sandwich ELISA Wells coated with purified
antibody Add sample to be tested for
antigen Wash of unbound antigen Add enzyme labeled specific
antibody Wash off unbound antibody “+” control → known antigen “-” control → omit known
antigen
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Tracking Down the Infection
A
B
C
D
E
F
G
+
-
H -
+
+
+
+
+
- + + +
+ + -
+ + + +
+ + + +
+
+
-
-
After running ELISA, you will write a plus our minus beside your name.
You will also mark a plus or minus by the name of the partners you shared
Whoever has the most pluses is revealed as the early sources of infections
Limitation: source cannot be traced to a single individual.
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Questions?