(MC089)

(in vitro Antigen and Antibody Interaction)

..

(Cell Mediated Immunity, CMI) /
(Humoral Immunity, HI) (Humoral Immune Response,
HIR)
( specificity)
phagocyte


(tumor marker)

noncovalent bond
1. Electrostatic force (Coulombic force)

2. Hydrogen bonding force hydrophilic group -OH, -NH2 , COOH

3. Hydrophobic force non-polar
hydrophobic group
4. Van der Waals force external electron
clouds
4

(complementary) (lock and key)

pH: pH 7.2-7.4 pH

:
4-40 oC (denature)
Ionic strength: ion salt
0.15 M NaCl (0.9% NaCl , isotonic solution) physiological salt
solution normal saline solution (NSS)

5
1. (neutralization)
2. (precipitation)
3. (agglutination)
4. (complement dependent reaction)
5. (labeled immunoassay)

1. Neutralization
(neutralize)
( neutralizing
antibody) neutralizing antibody

neutralizing antibody
neutralizing antibody
streptolysin O
Streptococcus pyogenes
streptolysin O

streptolysin O, streptolysin O
streptolysin O, streptolysin O
anti-streptolysin O (ASO) test
Streptococcus pyogenes (-Streptococcus group A)
Rubella, Influenza, Dengue
(hemagglutinating
antigen) neutralization

neutralization hemagglutination inhibition

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc

2. Precipitation
(soluble antigen)
antigen-antibody complex
lattice formation
1 1
1
(lattice)
1
3
1. antibody excess zone (pro zone)
antigen-antibody complex
2. equivalent zone

3. antigen excess zone (post zone) antigenantibody complex soluble immune

complex (soluble antigen-antibody complex)

Supernate
Precipitate

free Ab

Absence of free
Ag + Ab

Free Ag

Precipitate
= Antigen
= Antibody

Ab excess

Equivalence

Ag excess

Increasing amount of antigen

1.
3

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc

2.1

(semisolid media) (gel) (diffusion)

2.1.1 Double immunodiffusion (Ouchterlony's method)

(diffuse)
(precipitin band, precipitin line)

( 2)

Ag

Ab

2. double immunodiffusion
(Ouchterlony's method)


3 3 3 3 3
Ag1

Ag2
1
Ab

3 .

3 .
3
Line of identity

3 .

3 .
6

Line of partial identity

Line of non-identity
3. double immunodiffusion
(Ouchterlony's method)
4

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc

 3 1 2
1 2 (identity)
3 6 4
4 5 4 6
5 6 4
(spur) 4 6
5 4 5
(partial identity)
3 7 8
(non-identity)

2.1.2 Single radial immunodiffusion, SRID (Mancini's method)

(diffuse)

( 4)

( 5)

diameter (d)

Antigen concentration

15

10

unknown

d2

Standard antigen (mg/ml)

4 .
SRID

5 .
(d2 )

2.2 electrophoresis
5

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc


electrophoresis
Immunoelectrophoresis (IEP), Counter-immunoelectrophoresis (CIE), Rocket immunoelectrophoresis

2.2.1 Immunoelectrophoresis (IEP)

(electrophoresis)

1.
(-)

(+)

Ag

2. (trough)
trough ( Ab)

3.
()

2.2.2 Counter-immunoelectrophoresis (CIE)

2

2

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc

(-)

(+)

A g

Ab

body fluids
HBsAg ( anti-HBsAg ) anti-HBsAg ( HBsAg )
bacterial antigen ( H.influenzae, Streptococcus)

3. Agglutination
(particulate antigen)
(particle) (
)
(agglutination) agglutination

3.1 Direct Active agglutination

3.2 Indirect Passive agglutination

soluble antigen
(latex)
3.3 Reverse passive agglutination

soluble antigen
agglutination hem- passive
hemagglutination, reverse passive hemagglutination

4. Complement dependent reaction

7

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc

 Complement fixation test (CF)

2 test system
indicator system

antigen-antibody complex (fix)
indicator system (
antibody (sensitized cell))
sensitized cell
indicator system test system
indicator system

SRBC

Ag

SRBC

Ag

C'

C'C'

Ab

Ab

Ab

Test system

Indicator system

Test system

Indicator system

With Ab

No lysis

No Ab

lysis

fixes complement
5. Labeled Immunoassay

no C' fixation

(sensitive)
(labeling)
(fluorescent dye)
(enzyme) (isotope, radioactive substance)
Immunofluorescence (IF)

( )

(fluorescent microscope)

Enzyme immunoassay (EIA)

8

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc

 Enzyme-linked immunosorbent
assay (ELISA) immunofluorescence substrate

Radioimmunoassay (RIA)
(isotope) 2

F***

E***

R***

Labelled Ab

Speciific Ag
Immunofluorescence

Enzyme immunoassay

Radioimmunoassay

(titer, titre)
()
1:2, 1:4, 1:8 ... 1:16
1:32 1:16

1:16 16

1. . .
. 4. : .. , 2537: 159-189.
2. Sell S. Basic immunology: immune mechanisms in health and disease. New York : Elsevier, 1987:
117-148.
3. Zmijewski CM, Bellanti JA. Antigen-antibody interactions. Bellanti JA ed. Immunology III. Tokyo:
Igaku-Shoin/ Saunders, 1985: 160-175.

Ag-Ab Rx /Dr.Suwin Wongwajana/13 Oct.'99/newAg-Ab.doc