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COMPLEMENT

FIXATION TEST
(CFT)
Microbiology Pharmacy

Dhean Vetra

260110110151

Nafilah Najla

260110110152

Karina Novitasari 260110110153


Shintya Noor

260110110154

Alhamzah Rahmat

260110110155

Faisal F Fasa

260110110156

Fauzan Fikri

260110110157

Gita Gracia M 260110110158


Melati Nur A

260110110159

Ariqa Dhianur S
Nusaibah Al Hima
Aisya Amalia M

260110110160
260110110161
260110110162

Derisha Amalia P 260110110163


Casuarina R

260110110164

Melanie Wibowo

260110110165

Aditya Salman H 260110110166

Introductio
n

Result and
Interpretati
on

Principles

CF
T

Negative
Test

Component
s

Positive
Test

1. INTRODUCTION
The complement fixation test (CFT) was extensively used in syphilis
serology after being introduced by Wasserman in 1909.
1. Complement is a
protein (globulin)
present in normal
serum.

2. Whole
complement system
is made up of nine
components: C1 to
C9

4. Complement
binds to Ag-Ab
complex

3. Complement
proteins are heat
labile and are
destroyed by
heating at 56C for
20 30 minutes.

5. When the Ag is
an RBC it causes
lysis of RBCs.

2. PRINCIPLE
Complement takes part in many of the immunological reactions. It
gets absorbed during the combination of antigens and antibody.
This property of antigenantibody complex to fix the complement
is used in complement fixation test for the identification of
specific antibodies.
The haemolytic system containing sheep erythrocytes (RBC) and
its corresponding antibody (amboceptor) is used as an indicator
which shows the utilization or availability of the complement.
If the complement is fixed then there will be no lysis of sheep
erythrocytes, thus denoting a positive test.
If the complement is available then there will be haemolysis which
is a property of complement, denoting a negative test.

The test
requires five
reagents
and is
carried out
in two steps.

3. COMPONENTS OF CFT

1. Test
System
Antigen
: It may be soluble
or
particulate.
Antibody
: Human serum
(May or may not contain
Antibody towards specific
Antigen)
Complement
: It is pooled
serum obtained from 4 to 5
guinea pigs. It should be fresh
or specially preserved as the
complement activity is heat
labile (stored at -30 C in small
fractions). The complement
activity should be initially
standardized before using in the
test.

2.
Indicator
System
(Haemol
ytic
system)
Erythrocytes
: Sheep
RBC
Amboceptor(
Hemolysin)
:Rabbit
antibody to
sheep red cells
prepared by
inoculating
sheep
erythrocytes
into rabbit
under standard
immunization
protocol.

4. POSITIVE TEST
Step 1:
At 37C
Antigen + Antibody + Complement
gets fixed
(from serum)

Complement

1 Hour

Step 2:
At 37C
Fixed Complement complex + Haemolytic system
No Haemolysis
1 Hour

(Test Positive)

5. NEGATIVE TEST

Step 1:
At 37C

Antigen + Antibody absent + Complement


1 Hour

Complement not fixed

Step 2:

At 37C
Free Complement + Haemolytic system
Haemolysis
1 Hour (Test Negative)

6. RESULTS AND
INTERPRETATIONS:
No haemolysis is considered as a positive test.
haemolysis of erythrocytes indicative of a
negative test.
1
2
3
4
A

Microtiter plate showing Haemolysis (Well A3,


A3 and B4) and No Haemolysis (Well

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