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ANTIGEN- ANTIBODY REACTIONS-
PRECIPITATION REACTIONSby
Vinu P .V
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ANTIGEN ANTIBODY REACTIONS
Characterized as
Non-covalent interaction (similar to lock and key fit of
enzyme-substrate)
Bonds:
Hydrogen
Ionic
Hydrophobic interactions
Van der Waals forces
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PRECIPITATION REACTIONS -(Immunologic tests)
Soluble Ag + Ab optT&pH insoluble ppt
Normalsaline
Antibodies------Precipitins
Antigens -----Precipitinogen
Occurs in liquid media, or in gels (agar, agarose)
Ppt rxns ------highly sensitive for detection of antigens&
relatively less sensitive for detection of antibodies
Either qualitative or quantitative
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Precipitin curve ( Marrack hypothesis)
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Keeping Ab conc. fixed
pptn occurs only at the zone of equivalence
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Applications
Diagnosis of infections
Identify blood , semen stains etc in criminal investigations
Detection of food adulterants
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Types of ppt rxns
1. Ring test
2. Slide test
3. Tube test
4. Immunodiffusion/ gel precipitation Single diffusion in one dimension
Double diffusion in one dimension
Single diffusion in two dimension
Double diffusion in two dimension
5. Immunoelectrophoresis6. Electoimmunodiffusion
Counter current immuno electrophoresis (CIEP)
Rocket electrophoresis
Laurells electro immunodiffusion in two dimension
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Ring test
Antigen solution taken over column of antiserum in
a test tube ppt at the junction
Eg: CRP test , ascoli thrombo pptn test
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Slide test A drop each of antigen & antibody on a slide
Mixed by rotation Floccules
Eg: VDRL test for syphilis 8
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Tube test
Antigen soln & antiserum (patients serum) taken in a testtube mixed Incubated & observed for ppt
Eg : Kahn test for syphilis
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Immunodiffusion / Gel precipitation
PPtn rxns are allowed to occur in gel
Types
Single diffusion in one dimension
Double diffusion in one dimension
Single diffusion in two dimension
Double diffusion in two dimension
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Single diffusion in one dimension
(Oudin method)
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Double diffusion in one dimension(Oakley - Fulthrope procedure)
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Single diffusion in two dimension(Mancini method/ Radial immunodiffusion)
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Double diffusion in two dimension(Ouchterlony method) Molten agar poured in glass slides / petry dishes
Test soln of Ag & Ab diffuse frm the wells to form a ppt atthe equilence zone
If two ppt lines fuse ( V- shaped) identical Ags
Ppt lines fuse and project (Y- shaped) partial identity
Ppt lines cross each other non identity 14
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Immunoelectrophoresis
resolution
Used for analysis of Serum Protein &
immunoglobulin
Test-Agar in a slide
1. Well cut is filled with Ag mixture, electric currentis applied separation separate spots based
on electric charge on the proteins
2. Rectangular trough is cut parallel to the separation
and filled with Ab incubated
3. Arc shaped ppt bands, r better visualized by
staining
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Electoimmunodiffusion (Electro immuno assay)
Counter current immuno electrophoresis (CIEP)
Rocket electrophoresis
Laurells electro immunodiffusion in two dimension
Counter current immunoelectrophoresis
Detect Ag & Ab with opposite charge
Move towards each other , in electric field line of ppt
Eg: Detection of HBS Ag in serum, 16
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Rocket electrophoresis
Quantitativeestimation of human plasma antigens & viral
antigens
Ab is added to molten agar, poured on glass slide
Wells r cut & filled with antigen soln
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Current applied , allowed to diffuse, forms rocket like pptlines
Area of each antigen, its height initial conc of antigen 18
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Laurells electro immunodiffusion in two
dimension
Used to quantify each Ag in a mixture
Ag mixture is first electrophoritically separated into diff
components immunodiffusion
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REFERENCES
1. Ananthanarayanan and Panikerstext book of
microbiology, seventh edition, orient longman,page no:82-
84.
2. Prescott, Harley and Kleins Microbiology, page no:771-773.
3. Pharmaceutical Biotechnology by S.P Vyas , V. K . Dixith,
Page No 466-471
4. www.wikipedia.com
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