DEFINITIONS

Antigen • Evokes specific immune response – Specific

antibody/specifically sensitised T-cells• Any foreign substance can act as an antigen

Antibody • Synthesized in response to a foreign

substance• Antibodies – Globulins – Immunoglobulins

IgG – Protects the body fluids

IgA – Protects the body surface

IgM – Protects the blood stream

IgE – Mediates reaginic

hypersensitivity

Immunoglobulin roles..

Specific Antigen

Specific Antibod

y

Antigen-Antibody reaction

Types(Serological reactions)

• Precipitation

• Agglutination

• Complement fixation test

• Immunofluorescence

• ELISA

• Highly specific• Entire molecules react,not fragment• Both participates - formation of

precipitates & agglutinates

Characteristics..(Ag-Ab)

Ag-Ab

SurfaceFirm &

Reversible

(Ag-Ab) contd..

Affinity

Avidity

Intensity of attraction between Ag-Ab

Binding strength of Ag-Ab

Ab-----Bivalent-------

Ab

Ag—multivalent—Ag

• Initial interaction• Rapid,reversible,weakPrimary

• Change in Physical state• Leads to

agglutination,precipitation etc.,Secondary

• Invivo initiate chain reactions• Leads to tissue

damage,hypersensitivityTertiary

Stages..(Ag-Ab)

Invivo

• Ab mediated immunity-Infectious diseases• Tissue injury-Hypersensitivity&Autoimmune

diseases

Invitro

• Diagnosis of Infectious & Non infectious diseases

• Epidemiological surveys

General

• Detection• Quantitation

Importance..(Ag-Ab)

Antigens/Antibodies

• Ability of test to detect even very minute quantities of Ag/Ab

Sensitivity

• Ability of test to detect reaction between homologous Ags&Abs

Selectivity

Parameters of sero reactions

PrecipitationAgglutination

Precipitation..

Precipitinogen + Precipitin Insoluble precipitate (Soluble Ag) (Ab) (Ag-Ab)

ElectrolytespH:7.4/37*c

Precipitation

Suspended as

Floccules

Flocculation

Mech.of Precipitation..(Lattice Hypothesis)

Ab-----Bivalent-------Ab Ag—multivalent—Ag

MultiMolecul

arlattice

Opt

imum

pr

opor

tion

Zone Phenomenon

Prozone Postzone

Identification

--Human blood--Seminal fluid--Bacteria--Microbial components

Detection

--Antigen--Antibody

Standardization

--Toxins--Antitoxins

Apps..

CRP test

Ring Test

VDRL-Syphilis

Slide Flocculation test

Kahn test-Syphilis

Tube flocculation test

Techniques

• Gel>specific>>sensitive>Liquid• Gel- porosity-Ag+Ab- Precipitation band

Immunodiffusion Test

Distinct for diff.Antigens

1)Identity2)Cross reaction3)Non identity between different antigens

Indications

Types..Single diffusion in one dimension(Oudin’s procedure)

Double diffusion in one dimension(Oakley-Fulthrope procedure)

Use:To determine the no. of antigens

Single diffusion in two dimensions(Radial immunodiffusion)

Uses: For quantitation of Ags & Abs

Double diffusion in two dimensions (Ouchterlony Procedure)

Use : To compare 2 Ag-Ab systems. Diagnosis of bacterial,viral,fungal,parasitic diseases. Eg:Elek’s test-Diptheria

Immunoelectrophoresis

Use:Analysis of Human serum in diseases such as Myeloma,Heavy chain disease

Countercurrent Immunoelectrophoresis

Use : Used for detection of various Ags in body fluids Diagnosis of Infectious diseases such as meningococcal,streptococcal

Rocket electrophoresis(One dimensional single

electroimmunodiffusion)

Uses : Quantitation of Proteins and other Ags in various clinical problems

Two dimensional Immunoelectrophoresis(Laurell

’s Procedure)

Uses:To detect and quantitate human serum proteins To detect abnormal proteins in various body fluids

Agglutination..

Agglutinogen + Agglutinin Ag-Ab complex (ParticulateAg) (Ab) (Agglutination)

Nacl

pH:7.4/37*c

Mechanism same as Precipitation---LATTICE HYPOTHESIS

Use:Blood grouping&Cross matching

Use:Diagnosis of Typhoid,Typhus,Brucellosi

s

Agglutination..Types

• To detect Antibodies 1)Fail to Agglutinate corresponding Antigens 2)Inhibit Agglutination

Coomb’s Antiglobulin test

Incomplete/ Blocking Abs

Direct(Invivo) Indirect(Invitro)Hemolytic disease of new born(Rh incompatibility)

To demonstrate non agglutinating Abs in Brucellosis

Indirect Coomb’s test

PrecipitationAgglutination

Coating soluble Ag onto surface of

Carrier molecule

Instead of Ag-----Ab=Reverse Passive Agglutination

Passive Agglutination tests

Ag Adsorbed Latex(milk)

Latex Agglutination test

Specific Ab

Milky white liquid

Coarse suspensi

on of granules

Haemagglutination test

Eg:Treponema pallidium haemagglutination test for serodiagnosis of Syphilis

Coagglutination test

Uses:Detection of bacterial Ags in blood,CSF in pneumococcal,gonococcal infections etc.,

Tq…u…..

by S.Vigneshwaran