SEROLGY

Antigen and Antibody Reactions

Dr.T.V.Rao MD

Dr.T.V.Rao MD

Serology
The branch of laboratory medicine that studies blood serum for evidence of infection and other parameters by evaluating antigenantibody reactions in

vitro
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Serology
Serology is the
scientific study of blood serum. In practice, the term usually refers to the diagnostic identification of antibodies in the serum We can detect antigens too
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Types of Antigen and Antibody reactions

1. 2. 3. 4. 5. 6. 7. Agglutination tests Double diffusion precipitation tests Immunoelectrophoresis Western blot tests Complement fixation tests Immunofluorescence testing Immunoassays
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What happens in Antigen and Antibody reactions

React with each other in a observable manner. Uses 1 Helps antibody mediated immunity in infection, and tissue injury 2 Helps diagnosis of Infections. 3 In epidemiological surveys 4 Detections and quantization of antigens and antibodies
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Detectable reactions 2nd Stage

Reaction can occur as
1 Precipitation 2 Agglutination 3 Lysis and killing of live antigens 4Neutralizatiobn of toxins 5 Fixation of complement 6 Immobilization of motile microbes 7 Enhancement of Phagocytosis.
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General features of Antigen and antibody reactions.

Specific reaction combines with specific antigen Entire molecule reacts not fragments No denaturation of antigen or antibody Combination occurs as surface antigens to surface of antibodies Commination is firm but reversible depends on affinity and avidity Both antigens and antibodies participate Combine in varying proportions Bivalent and multivalent
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Terms used in evaluating test Methodology Sensitivity

Analytical Sensitivity ability of a
test to detect very small amounts of a substance

Clinical Sensitivity ability of test to

give positive result if patient has the disease (no false negative results)
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Specificity
Analytical Specificity ability of test to detect substance without interference from cross-reacting substances Clinical Specificity ability of test to give negative result if patient does not have disease (no false positive results)
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Affinity
Antibody affinity is the strength of the reaction between a single antigenic determinant and a single combining site on the antibody.
It is the sum of the attractive and repulsive forces operating between the antigenic determinant and the combining site .

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Avidity
Avidity is a measure of the overall strength of binding of an antigen with many antigenic determinants and multivalent antibodies Avidity is influenced by both the valence of the antibody and the valence of the antigen. Avidity is more than the sum of the individual affinities.
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Dilution
Estimating the antibody by determining the greatest degree to which the serum may be diluted without losing the power to given an observable effect in a mixture with specific antigen

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Measurement of Antigen and Antibody reactions

Measured as Mass Nitrogen (microgram) As Units As titer

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Titer
Different dilutions of serum are tested in mixture with a constant amount of antigen and greatest reacting dilution is taken as the measure or Titer

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Common methods in creating dilutions

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Expression of Titers
Expressed in term of the was in which they are made Dilution 1 in 8 is a dilution made by mixing one volume of serum with seven volumes of diluents
(Normal Saline )

Incorrect to express dilution as 1/8

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Majority Diagnostic tests are Serological tests

There are several serology techniques that can be used depending on the antibodies being studied. These include: ELISA, agglutination, precipitation, complement-fixation, and fluorescent antibodies.
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How Antigen Antibody reaction occurs

Primary stage Interaction between antigen and antibody occurs without any visible effects The reaction is rapid even at low temperature but the reaction is reversible Weaker _ intermolecular forces Van der walls force and ionic bonds Can be measured with radioactive isotopes and Florescent dyes.
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Ag-Ab interactions
Bonds:
Hydrogen Ionic Hydrophobic interactions Van der Waals forces

Each bond is weak; many are strong To hold they must be close requiring high amts of complementarity!

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What happen further ?

1 Precipitation 2 Lysis of cells. 3 Killing of live antigen 4 Neutralization of toxins 5 Complement fixation 6 Immobilization of motile microbes 7 Enhancement of Phagocytosis 8 Entire molecule react no fragmented 9 No denaturation 10 Combination occurs on surface 11 Firm but reversible
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Types of serological tests

1. 2. 3. 4. 5. 6. 7. Agglutination tests Double diffusion precipitation tests Immunoelectrophoresis Western blot tests Complement fixation tests Immunofluorescence testing Immunoassays
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Immunology/ Serology? Precipitation Reactions

Capillary tube precipitation (Ring Test) Ouchterlony Double Diffusion (Immunodiffusion) Radialimmunodiffusion (RID) Immunoelectrophoresis (IEP) Rocket Electroimmunodiffusion (EID) Counterimmunoelectrophoresis (CIEP)
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Antigen Antibody
reactions presenting with precipitation

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Precipitation reactions
Precipitation- classic demonstration of antibody-antigen interaction Antibody and soluble antigen aggregate to form a visible precipitate Antibody must be bivalent (Fabs wont work) Antigen must be multivalent
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Precipitation - Reaction
In precipitation antigen combines with its antibody in the presence of electrolytes ( Nacl ) at a suitable temperature and Ph the antigen and antibody complexes form a insoluble precipitate suspended as floccules. Reaction can take place in liquid medium, gels, agar, agarose, polyacrylamide
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Precipitation
Principle
Soluble antigen + antibody (in proper proportions) > visible precipitate Lattice formation (antigen binds with Fab sites of 2 antibodies) Double diffusion (Ouchterlony) Single diffusion (radial Immunodiffusion) Immunoelectrophoresis Immunofixation
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Examples

How precipitation occurs

Precipitation occurs with most antigens because the antigen is multivalent (i.e. has several antigenic determinants per molecule to which antibodies can bind). Antibodies have at least two antigen binding sites (and in the case of IgM there is a multimeric complex with up to 10 antigen binding sites), thus large aggregates or gel-like lattices of antigen and antibody are formed Specific reaction Antigen combines with homologusantibody - vice versa

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Mechanism of Precipitation
Lattice hypothesis Multivalent antigens combine with bivalent antibodies in varying proportions depending on the antigen and antibody ratio in the reacting mixture

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ANTIGEN EXCESS

ZONE of Equivalence No Soluble Ag or Ab

ANTIBODY EXCESS

Ab CONC

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Precipitation Reactions

( no precipitate is formed ( Lattices or if an Ag contains only a large aggregates ) single copy of each epitope ) FIGURE 6-4

Precipitation reactions in fluids yield a precipitin curve.

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Applications of precipitation reactions

Qualitative and quantitative More useful for antigen detection to as least as 1 microgram Blood, seminal stains, Food adulteration
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Slide test -Flocculation test VDRL test

VDRL Test a drop of VDRL antigen to a drop of patients serum, Shake The reaction observed under microscope Observe for flocculation reaction A Khan test is done in a test tube

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Non reactive and Reactive VDRL Tests

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Agglutination+: ve

RPR Agglutination

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Tube test - Precipitation

Kahn test for Syphilis a tube flocculation test. Quantitative tube flocculation test used in standardarisation of toxin/toxoid Serum dilution of toxin or toxoid is added to tubes containing a fixed quantity of antitoxin The amount of toxin that flocculates optimally with one unit of antitoxin is defined as Lf dose
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Single diffusion in one Direction

O
Oudin Procedure

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Immuno Diffusion Tests

Can demonstrate the immunological identity (or not) of two antigen samples; radial
Immunodiffusion
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Ring Test
The reaction is demonstrated by layering antigen solution over the column of antigen in a narrow tube, The precipitate forms at the junction of two liquids. Eg Ascolis thermo precipitate test Grouping of streptococci C-reactive protein

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Oudin Procedure
Antibodies in agar gel Above antigen is layered Single diffusion in one direction Called Qudin procedure Antibody is incorporated in agar Antigen diffuses down Precipitation concentration of antigen at the site increases due to diffusion
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Double diffusion in one dimension Oakley Fulthrope procedure

Antibodies in gel Agar layer Antigen layered Antigens and antibodies moves towards each other Forms precipitate
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Single Diffusion In two Dimensions Radial Immunodiffusion

Radial Immunodiffusion is an Immunodiffusion technique used in immunology to detect quantity of antigen by measuring the radius surrounding samples of the antigen, marking the boundary between it and antibody
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DOUBLE DIFFUSION

Antigen

Antibody

Immune Complex
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Antigen

Antibody

Antigen

Immune Complexes

Zone of Equivalence
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OUCHTERLONY ANALYSIS Diffusion of Antigens and Polyclonal Antibodies

Non-Identity
Antigen 1
(Molecule #1)

Antigen 2
(Molecule #2)

Antibodies to both antigens

The same Animal was injected with
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OUCHTERLONY ANALYSIS

Partial - Identity

This animal was only injected with Antigen #4

Antigen 3
is a part of antigen 4

Antigen 4

Also remember that this antibody is a multi-clonal antibody such as an antiserum to an antigenic

Antibody
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Remember that Protein Antigens have different antigenic 46 determinants

Single Diffusion In two Dimensions Radial Immunodiffusion

Antigen is added to the wells cut on the surface of the gel It diffuses radially from well and forms a ring shaped band of precipitation The halo of precipitation diameter gives the estimate of concentration of antigen Used in estimation of Immunoglobulin's
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Precipitation in Agar
These tests are done in agar Tested for passive immuno diffusion in Agarose There are several methods in testing this procedure
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Eleks Immunodiffusion Test

Elek Immunodiffusion test. Sterile filter paper impregnated with diphtheria antitoxin is imbedded in agar culture medium. Isolates of
C diphtheria are then streaked across the plate at an angle of 90 to the antitoxin strip. Toxigenic C diphtheria is detected because secreted toxin diffuses from the area of growth and reacts with antitoxin to form lines of precipitin

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Radial Immunodiffusion
Radial Immunodiffusion, a variation of the agar precipitation technique, is used in clinical immunology for the detection and quantitation of all classes of Immunoglobulin's, complement, ceroplastic, transferring, and other serum components
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Double Diffusion in one Dimension Oakley Fulthrope procedure

In Double diffusion in one direction antibody in agar gel moves through the layer of plain agar to the antigen above They react to each other Form a band of precipitation Virus antigen is placed in the central well and diffuses outwards. Wells A and C contain positive sera, well B contains a negative sample. The black areas show where antibody in the positive sera have bound to virus antigen and formed a precipitate

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Double diffusion in two dimensions Ouchterlony procedure

Both antigen and antibody diffuses independently through agar gel in two dimensions horizontally and vertically Done in a slide or petridish In 12 48 hours line of precipitates are formed Useful in serological identification

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Immunoelectrophoresis:
In Immunoelectrophoresis, a complex mixture of antigens is placed in a well punched out of an agar gel and the antigens are electrophoresed so that the antigen are separated according to their charge. After electrophoresis, a trough is cut in the gel and antibodies are added. As the antibodies diffuse into the agar, precipitin lines are produced in the equivalence zone when an antigen/antibody reaction occurs.
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Immuno-electrophoresis
Mixtures containing multiple antigen species which cross react with the same antiserum may be analysed by running them first on an analytical gel, then cutting a strip from that gel and laying it in a slit cut into the immunoelectrophoresis gel to form a large well. The result is a pattern which shows the positions of strongly reacting antigen species.
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Immunoelectrophoresis
Immunoelectrophoresis --migration of molecules due to electric charge Positive particles travel to cathode Negative particles travel anode Precipitin specificity provides a critical indicator of identity
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Immuno Electrophoresis
In this procedure the electrophoretic separation of compatible antigen into constituent protein followed by Immunodiffusion against its antiserum resulting in separate precipitation line, indicating relation between each individual protein in the antibody

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Electrical gel Immunoelectrophoresis

The reaction is electrically driven, different antigens are separated according to their charges under electrical fields Number of antigens can be identified
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Countercurrent electrophoresis
Method
Ag and Ab migrate toward each other by electrophoresis Used only when Ag and Ab have opposite charges

Ag Ab

Qualitative
Rapid
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Counter current Immunoelectrophoresis

In this procedure movement of antigens towards the anode and antibody towards the cathode through agar under electric field Useful studies on CSF Hepatitis B surface antigen detection Detection of fetoproteins

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Imunoelectrphoresis (IEP) Qualitative

A serum sample is electrophoresed through an agar medium. A trough is cut in the agar and filled with Ab. A precipitin arc is then formed. Because Ag diffuses radially and Ab from a trough diffuses, the reactants meet in optimal proportions for precipitation.
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Rocket electrophoresis Laurel

The reactions appear as rocket Driven by electric current Rocket ImmunoElectrophoresis is used as a rapid way to quantitate antigen in complex samples.
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Immuno-electrophoresis gel
Mixtures containing multiple antigen species which cross react with the same antiserum may be analyzed by running them first on an analytical gel, then cutting a strip from that gel and laying it in a slit cut into the immunoelectrophoresis gel to form a large well. The result is a pattern which shows the positions of strongly reacting antigen species.
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Counterimmunoelectrophoresis

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Electro Immunodiffusion

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Rocket electrophoresis
Crossed Immunoelectrophoresis of antigens and antiserum. In the first dimension, proteins are separated by standard electrophoresis. The separated proteins are then run into the second dimension gel at an angle of 90 from the first dimension.

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Serology can be done on various specimens

Some serological tests are not limited to blood serum, but can also be performed on other bodily fluids such as semen and saliva, which have (roughly) similar properties to serum. Serological tests may also be used forensically, generally to link a perpetrator to a piece of evidence (e.g., linking a rapist to a semen sample).
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Measurement of Precipitation by Light

Antigen-antibody complexes, when formed at a high rate, will precipitate out of a solution resulting in a turbid or cloudy appearance. Turbidimetry measures the turbidity or cloudiness of a solution by measuring amount of light directly passing through a solution.
Nephelometry indirect measurement, measures amount of light scattered by the antigen-antibody complexes.
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Measurement of Precipitation by Light

Antigen-antibody complexes, when formed at a high rate, will precipitate out of a solution resulting in a turbid or cloudy appearance. Turbidimetry measures the turbidity or cloudiness of a solution by measuring amount of light directly passing through a solution.
Nephelometry indirect measurement, measures amount of light scattered by the antigen-antibody complexes

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Agglutination

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Agglutination
is the aggregation of particulate matter caused by the combination with specific antibody 1896: First observed by Gruber and Durham when serum antibody was found to react with bacterial cells
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Agglutination
Agglutinins
Antibodies that produce such reactions

Involves two-step process:

Sensitization or initial binding Lattice formation or formation of large aggregates
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Agglutination
Types of particles that participate in such reactions:
Erythrocytes Bacterial cells Inert carriers such as latex particles
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Secondary phenomenon: LATTICE FORMATION

Ab + multivalent Ag stable network (visible reaction) conc. of Ag and Ab Governed by physiochemical factors:
Ionic strength of milieu pH temperature

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Secondary Phenomenon
Lattice Formation The Fab portion of the Ig molecule attaches to antigens on 2 adjacent cells-visible results in agglutination If both antigen and antibody are SOLUBLE reaction will become visible over time, ie, precipitation

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Tube Agglutination Test

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Tube Agglutination Test

Agglutination No agglutination

1/10

1/20

1/40

1/80

1/160

1/320

Neg. ctrl

In this case, the titre is 1/40

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DIRECT AGGLUTINATION
-Test patient serum against large,

cellular antigens to screen for the presence of antibodies. Antigen is naturally present on the surface of the cells. In this case, the Ag-Ab reaction forms an agglutination, which is directly visible.
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Slide Agglutination Test

Used for serotyping (e.g. Salmonella) Antigen: isolated Salmonella in suspension Antibody: specific antisera against Salmonella Place test Salmonella in a drop of saline on a slide Add a drop of antiserum, mix and rock slide for approx 1 minute Examine for agglutination
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Slide Agglutination Test

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Agglutination Test

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OTHER DIRECT AGGLUTINATION TESTS

The particle antigen may be a bacterium. e.g.: Serotyping of E. coli, Salmonella using a specific antiserum The particle antigen may be a parasite. e.g.: Serodiagnosis of Toxoplasmosis The particle antigen may be a red blood cell. e.g.: Determination of blood groups
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Quantitative Micro Hemagglutination Test (HA)

Haemagglutination Tests
(HA)
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HEMAGGLUTINATION
Detects antibody to erythrocyte antigens sufficient concentration of antibody present-> antibody cross-link= agglutination non-reactive/insufficient antibody present= no agglutination

Binding different antigens on the RBC surface = detect antibodies to antigen other than those present in the cells
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HEMAGGLUTINATION
Chromic chloride, tannic acid, and glutaraldehyde= cross- link antigens to the cell IgG (does not agglutinate directly)-> need enhancement medium-> AHG AHG binds to the second antibody present on the erythrocyte.
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Haemagglutination

RBC

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Viral Haemagglutination
Some viruses and microbes contain proteins which bind to erythrocytes (red blood cells) causing them to clump together
NDV Adenovirus III AIV IBV Mycoplasma

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Hemagglutination test: method

1:8 1:2 1:2 1:2 1:2 1:2 virus

serial dilution 8 mix with red blood cells side view 16 32 64 128 256

top view

Titer = 32 HA units/ml

One HA unit :minimum amount of virus that causes Dr.T.V.Rao MD 89 complete agglutination of RBCs

HEMAGGLUTINATION INHIBITION TEST (HI)

VIRUSE
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SERUM
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In the absence of anti-virus antibodies

Erythrocytes

Virus Virus agglutination of erythrocytes

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In the presence of anti-virus antibodies

Erythrocytes

Virus

Anti-virus antibodies Viruses unable to bind to the erythrocytes

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Reading/Grading Agglutination Reactions

Done by gently shaking the tubes containing the serum and cells, and observing the cell button as it is dispersed Hard shaking must be avoided because this may yield to false result Attention should also be given to whether discoloration of the supernatant is present (Haemolysis).
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Antibody Titer
Is the lowest concentratio n of antibodies against a particular antigen.
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Coombs (Antiglobulin)Tests
Applications
Detection of anti-Rh Ab

Autoimmune hemolytic anemia

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Coombs (Antiglobulin)Tests
Incomplete Ab Direct Coombs Test Detects antibodies on erythrocytes

+
Patients RBCs Coombs Reagent (Antiglobulin)
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Coombs (Antiglobulin)Tests
Indirect Coombs Test
Detects anti-erythrocyte antibodies in serum
Step 1 Patients Serum Step 2

+
Target RBCs

Coombs Reagent (Antiglobulin) Dr.T.V.Rao MD

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Passive Agglutination
An agglutination reaction that employs particles that are coated with antigens not normally found in the cell surfaces Particle carriers include:
Red blood cells Polystyrene latex Bentonite charcoal
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Passive Agglutination
Passive agglutination has been used in the detection of :
Rheumatoid factor Antinuclear antibody in LE Ab to group A streptococcus antigens Ab to Trichinella spiralis
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Reverse Passive Agglutination

Antibody rather than antigen is attached to a carrier particle
For the detection of microbial antigens such as: Group A and B streptococcus Staphylococcus aureus Neisseria meningitidis Haemophilus influenzae Rotavirus Cryptococcus neoformans Mycoplasma pneumoniae Candida albicans
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Coagglutination
Name given to systems using inert bacteria as the inert particles to which the antibody is attached S.aureus: most frequently used because it has protein A in its outer surface that naturally adsorbs the Fc portion of the antibody
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 Highly specific but not very sensitive in detecting small quantities of antigen

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False-Positive Result
If injected with hCG to trigger ovulation or to lengthen luteal phase of menstrual cycle. Chorioepithelioma, hydatidiform mole or ingestion of aspirin To detect the presence of a testicular tumor in men False Negative Testing before reaching detectable levels of hCG.
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Immunofluorescence
Antibodies can be labeled with fluorescent dye Can localize binding sites on cell Dyes: Fluorescein, rhodamine, phycoerythrin can be conjugated to Fc region of Ab (so antigen binding is unaffected Absorb at one wavelength and emit at another
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Fluorescence

UV Light

Antigens on Cells or on Tissue Sections

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Fluorescence

Double layer Sandwich

UV Light

Antigens
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Enzyme Immunoassay ( EIA )

Introduced in 1966 alternative to fluorescent methods Versatile, simple economical Absence of radiation. EIA means measuring enzymes labelled antigen, hapten, antibody
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ELISA
Enzyme Linked Immuno-Sorbant Assay
Peroxidase Enzyme is permanently attached to Antibody Probe
Substrate that turns from clear to green

Ag

Ag

Microtiter ELISA
Antigens are immobilized toMD plastic surface of a the Dr.T.V.Rao Microtiter Plate
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Fluorescence

UV Light

Antigens on Cells or on Tissue Sections

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ELISA
Enzyme Linked Immuno-Sorbant Assay
Peroxidase Enzyme is permanently attached to the Antibody Probe

Substrate that turns from clear to green Ag Ag

Microtiter ELISA
Antigens are immobilized toMD plastic surface of a the Dr.T.V.Rao Microtiter Plate
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ELISA
The ELISA (Enzyme-Linked ImmunoSorbant Assay) can be used both qualitatively and quantitatively to measure antigen-antibody binding. Depending on what variation you use, it will detect antigen (hormones, enzymes, microbial antigens, illicit drugs) or antibody (anti-HIV in the screening test for HIV infection) in body fluids or tissue culture supernatants.
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ELISA
Enzyme-Linked Immuno-Sorbant Assay, also called ELISA, Enzyme
ImmunoAssay or EIA, is a biochemical technique used mainly in immunology to detect the presence of an antibody or an antigen in a sample. The ELISA has been used as a diagnostic tool in medicine and plant pathology, as well as a quality control check in various industries.
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Enzyme Linked Immuno Assay

The Technique involves use of Immuno-Sorbant and absorbing material specific for one of the components of reaction, the antigen or antibody
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Components in ELISA testing

Conjugate Horseradish peroxidase Substrate - O-phenyle diamine dihydrochloride The test is conducted in solid phase Polystyrene, Polyvinyl or polycarbonate tubes or in plastics
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ELISA plate

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ELISA methodology
Performing an ELISA involves at least one antibody with specificity for a particular antigen. The sample with an unknown amount of antigen is immobilized on a solid support (usually a polystyrene microtiter plate) either non-specifically (via adsorption to the surface) or specifically (via capture by another antibody specific to the same antigen, in a "sandwich" ELISA).
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ELISA Methodology

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ELISA methodology
After the antigen is immobilized the detection antibody is added, forming a complex with the antigen. The detection antibody can be covalently linked to an enzyme, or can itself be detected by a secondary antibody which is linked to an enzyme through bioconjugation
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Sandwich ELISA

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Different methods of ELISA

Direct and Indirect ELISA Sandwich Non competitive Sandwich
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ELISA most popularly used method

The ELISA is probably the most commonly used immunological assay because of its versatility, sensitivity (ability to detect small amounts of antigen or antibody), specificity (ability to discriminate between closely related but antigenically different molecules), and ease of automation. Although some of the substrates are carcinogenic, they are generally considered safer than radioisotopes used in RIA (radioimmunoassay).
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Uses of ELISA
Helps detection of Antigens, Antibodies, hormones and Enzymes Eg in Microbiology Antigens HbS Ag Antibodies HIV, HCV, CMV, several other disease
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Radio Immuno Assay Berson and Yallow

Besides fluorescent dyes other labels can be used Uses with Radio isotopes Variety of tests are done for detection of antigen or antibody The term binder ligand assay has been used The minute amounts of substances can be detected Used in Biology and Medicine
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Rosalyn S. Yalow and Sol Berson

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RIA ( Radio Immuno Assay )

The technique was introduced in 1960 by Berson and Yalow as an assay for the concentration of insulin in plasma. It represented the first time that hormone levels in the blood could be detected by an in vitro assay.
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Uses of RIA
Used for detection of Hormones, enzymes,tumour markers IgE and viral antigens RIA is a competitive binding assay fixed amount of antibody and radiolabelled antigen react in the presence of unlabelled antigen Detection is done for free and bound fractions, ratios calculated
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Immunofluorescence
The purpose of immunofluorescence is to detect the location and relative abundance of any protein for which you have an antibody. Once you have antibodies to your favourite protein, you can use them to indicate where the protein is located.
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Immunofluorescence

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Fluorescent Methods

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Direct and Indirect Methods

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Western Blot
Western blot analysis can detect your protein of interest from a mixture of a great number of proteins. Western blotting can give you information about the size of your protein (with comparison to a size marker or ladder in kDa), and also give you information on protein expression (with comparison to a control such as untreated sample or another cell type or tissue).
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Western Blot Test

The method originated from the laboratory of George Stark at Stanford. The name western blot was given to the technique by W. Neal Burnette and is a play on the name Southern blot, a technique for DNA detection developed earlier by Edwin Southern. Detection of RNA is termed northern blotting.
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Appearance of test readings

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Flowcytometry
FACS- fluorescence-activated cell sorter Analyze cell populations Sort cells with different features into different containers (e.g., T and B cells; cells that are producing a cellsurface marker from those that are not)
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Uses for flow cytometry

Percentage of a total population of cells Measuring antigen density within a population of cells Multiple antibodies can be used to assess several cell surface antigens simultaneously Clinical analysis (tumor characterization)
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Chemiluminescence's
Chemiluminescence's Chemical reaction emitting energy in the form of light Chemilumiscence - Luminol or acridinium esters causes signal in the process of antigen antibody reaction Signal can be amplified, measured, and the concentration of the analyses sample Uses the automated methods. Increasingly used where the volume of work is large

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Chemiluminescence's Immuno Assay

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Immunochromatographic Tests
One step in diagnosing Simple Economical. Reliable Eg HbsAg A small cassette system containing a membrane impregnated with antiHbsAg antibody colloidal gold dye conjugate The membrane is exposed at three windows on the cassette
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Immunochromatographic Tests

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Immunochromatographic Tests
A colored band appears at the second window Control also can be recorded
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Also called as Dot Methods

The tests can be done by paramedical staff, as they are simple to read

Helps in emergency rooms. The results are available within few minutes The HIV and HBV infections can be done at the earliest
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Results can be read as Positive and Negative at the earliest

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 Programme created by Dr.T.V.Rao MD for Basic learning in Immunology

Email doctortvrao@gmail.com
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