Neutralization

Neutralization
and Complement
and Complement
Fixation
Fixation

LACHICA, KRISTIN JANE MARIZ G.

ONG, JESSICA MAE L.
REYES, CHERRY ROSE T.
SALVADOR, PAMELA MARIE M.
TAN, BERNADETTE IRISH T.
NEUTRALIZATION
Antigen-antibody reaction in which
the reactive effect of a particular
antigen is nullified by a specific
antibody.

POSITIVE: NEGATIVE:

No Agglutination Agglutination

The reaction is used to identify toxins and antitoxins, as

well as viruses and viral antibodies.
TOXIN NEUTRALIZATION
In-Vitro Assay (ASO)

Streptolysin O rgt added to serial dilution of

●
patient serum (infected with B- Streptococci)

ASO titer = highest dilution w/o hemolysis

Todd units
Schick Test
 In vivo
 It is used to test for immunity to diptheria

Diptheria
Toxin
Antitoxin antibodies

YES NO

NO REACTION SKIN BECOMES RED AND

- SWOLLEN +

Similar: Dicks test = Scarlet fever

Application of Tests In-Vivo

Protection test
●
Assess the potency of a
vaccine
●
In vivo procedure
Application of Tests In-Vivo

Standard toxin Test serum

Application of Tests In-Vivo

If neutralizing antibody is present, toxin will be neutralized.

Animal will show no sign of infection
Active Assay
 Immunize the animal with a particular vaccine
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Animal develops
Introducing a of
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 Active Assay
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Produceorany
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 LD50 (Lethal dose)

 Lowest dose of the microbes that will kill 50% of
lab. animal
Passive Assay

Collect serum sample of different dilutions

Add standard amount of microbial suspension

Determine the highest concentration that will protect the

animal
Passive Assay

ED50
●
Effective dose
●
Completely neutralize the
pathologic effect
 Viral Neutralization
Loss of infectivity
through
Loss reaction of the
of infectivity
virus with
through specificof the
reaction
antibody
virus with specific
antibody

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Viral Neutralization

In-vivo test

In-ovo test

Pock Reduction Test

Plaque Reduction Test

Cytopathic Effect
Viral Neutralization

In-Vivo Test

●
Uses live animals as in animal protection test

In-Ovo Test

●
Uses chorioallantoic membrane (CAM) of embryonated chicken egg for the
inoculation of virus-antibody mixture.
Viral Neutralization

In Ovo Test
Viral Neutralization

Pock Reduction Test

●
Neutralizing antibodies in serum of virally infected patients neutralize the virus
leading to reduction in number of pocks in embryonated egg.

Plaque Reduction Test

●
Similar to Pock Reduction Test
●
Reduction in the number of plaques (flat area) in tissue culture
Viral Neutralization

Cytopathic Effect

●
Neutralizing antibodies in serum of virally infected patients prevent the virus
from altering the tissue culture (cytopathic effect or pathologic changes).
Complement Fixation

Used to detect for the presence of specific antibody or antigen in a patient’s

serum.

Diagnosis of many bacterial, viral and other diseases, including syphilis.

2 Stages of C’ Fixation:

Stage 2: detection of residual

Stage 1: binding or fixation of
unbound complement, by its
complement if certain antigen-
hemolytic action after the sensitized
antibody reaction occur
RBC is added
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
Complement Fixation
PRINCIPLE

First reaction involves the reaction

between the antigen and antibody
forming the antigen- antibody
complexes, and in the process the
complement is “fixed” by the
complex
PRINCIPLE

Second reaction involves the testing

for the presence of “free”
complement or “unfixed”
complement in the unknown serum.
This is done by adding sensitized
RBC sensitized with specific
hemolysin
Interpretation
Thank you!