Antigen Antibody Reaction And Precipitation Reaction
Antigen antibody reaction
- Are reversible specific non-covalent biochemical reactions
- It is a specific chemical interaction between antibodies produced by B cells of the white blood cells and antigens during immune reaction.
- Precipitation reactions are based on the interaction of antibodies and antigens.
- They are based on two soluble reactants that come together to make one insoluble product, the precipitate
- Form basis for
- Humoral immunity or antibody mediated immunity
- Detection of infectious disease causing organism
Ag-Ab reaction occurs in three stages
Primary Stage
- Formation of Ag-Ab complex
- Ag-Ab combined by weaker intermolecular forces
- van der Waals,
- ionic bond
- H2 binding
Secondary stage
- leads precipitation
- agglutination
- lysis of cells etc.
Tertiary stage (reaction):
- Leads to tissue damage
- Destruction of Ag or its Neutralization
- Types of Antigen-antibody reactions
Types of antigen- antibody reactions in vivo:
- Agglutination
- Precipitation
- Complement fixation
- Neutralization
- Antibody dependent cell mediated cytotoxicity (ADCC)
- Immobilization
Types of antigen antibody reactions used in vitro:
- Agglutination
- Precipitation
- Neutralization
- Complement fixation
- Fluorescent Fluorescent-antibody technique antibody technique
- ELISA- Enzyme-linked immunosorbent assay
- Radioimmunoassay
- Immunochromatography (ICT)
Precipitation reaction
- Sensitive in detection of antigens
- As little as 1 g of protein can be detected by precipitation tests.
- Precipitation is relatively less sensitive for the detection qf antibodies.
Mechanism of precipitation.
- Marrack (1934) proposed the lattice hypothesis
- Based on the interaction of antibodies and antigens.
- Two soluble reactants that come together to make one insoluble product, the precipitate .
- When antigen and antibody exist in optimal proportions formation of lattices (cross-links).
Method
- Soluble antigen combines with its antibody in the presence of electrolytes (NaCI)
- At a suitable temperature and pH
- Antigen antibody complex forms an isoluble precipitate.
The amount of precipitate formed is greatly influenced by the:
- Relative proportions of antigens and antibodies
- The Ab avidity
Plot in a curve
- Three zones are detected ,If
- Varying amounts of Ag are mixed and incubated with Constant volume of antisera
Zone of Ag excess :
- insufficient Ab
- Too small complexes to precipitate
Equivalence zone :
- Large lattice is formed
- Visible precipitates
Zone of Ab excess :
- not enough Ag
- Too small complexes to precipitate.
Types
Ring test
- Simplest type
- e.g. Ascoli’s thermoprecipitin test and streptococcal lancefield grouping.
Slide test
- VDRL test of syphilis
- When the heat inactivated (to destroy complement) serum of patient is reacted with freshly prepared non-treponemal antigen, flocculation reaction (antigen and antibody complex are suspended) occurs.
- The flocculation can be observed by using microscope
Tube test
- Kahn test of syphilis
Immunodiffusion/Counter Current immunoelectrophoresis
- ppt in gel
- e.g. Elek test for toxigenicity in diphtheria (double diffusion).
Electroimmunodiffusion
- e.g. rocket electrophoresis for quantitative estimation of antigen.
Exam Question
Antigen antibody reaction
Primary Stage
- Formation of Ag-Ab complex
- Ag-Ab combined by weaker intermolecular forces
- van der Waals,
- ionic bond
- H2 binding
Secondary stage
- agglutination
Tertiary stage (reaction):
- Leads to tissue damage
Precipitation reaction
Mechanism of precipitation.
- Marrack (1934) proposed the lattice hypothesis
- When antigen and antibody exist in optimal proportions formation of lattices (cross-links).
Method
- Soluble antigen combines with its antibody in the presence of electrolytes (NaCI)
- At a suitable temperature and pH
- Antigen antibody complex forms an isoluble precipitate.
Plot in a curve
- Three zones are detected ,If
- Varying amounts of Ag are mixed and incubated with Constant volume of antisera
Zone of Ag excess :
- insufficient Ab
- Too small complexes to precipitate
Equivalence zone :
- Large lattice is formed
- Visible precipitates
Zone of Ab excess :
- not enough Ag
- Too small complexes to precipitate.
Types
Ring test
- e.g. Ascoli’s thermoprecipitin test and streptococcal lancefield grouping.
Slide flocculation test
- VDRL test of syphilis
- Antigen and antibody complex are suspended..
- The flocculation can be observed by using microscope
Tube test
- Kahn test of syphilis
Immunodiffusion/Counter Current immunoelectrophoresis
- e.g. Elek test for toxigenicity in diphtheria (double diffusion).
Electroimmunodiffusion
- e.g. rocket electrophoresis for quantitative estimation of antigen.
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