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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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