- Primary serological tests: (Marker techniques) e.g.

Enzyme linked immuonosorbent assay (ELISA)Immuno flurescent antibody technique (IFAT)Radio immuno assay (RIA)

- Secondary serological tests: e.g.

Agglutination testsComplement fixation tests (CFT)Precipitation testsSerum neutralization tests (SNT)Toxin-antitoxin test

Classification of antigen-antibody interactions:

-Antigens are whole cells (eg. RBC or bacteria).

-Antibodies cross-link the antigens.

-Types of tests:

-Rapid plasma test (syphilis). -Cold agglutinin test (mycoplasma).

-Weil-Felix test (rickettsial). -Latex agglutination test (pregnancy).

Agglutination:

-Agglutination reactions involve whole cell antigens, while precipitation reactions involve soluble antigens.

-Cellular/molecular view of agglutination and

Precipitation reactions that produce visible Ag-Ab complexes.

Agglutination : ( continued )

Agglutination : ( continued )

-The rapid plasma reagin (RPR) test is available as a kit .

The patient's serum is mixed with cardiolipin on a plastic-coated card that can be examined with the naked eye.

Rapid plasma reagin ( RPR test )

Precipitation

-Soluble antigens.

-Binding of antibody to antigen, makes the complex insoluble or visible.

-Types of test:

-Tube precipitation .

-Ouchterlony (double diffusion).

-Immunoelectrophoresis.

Precipitation : ( Continued )

-Precipitation reactions

Precipitation : ( Continued )

Ouchterlony (double diffusion)

Precipitation : ( Continued )

Precipitation curve :

Complement fixation

-Antibody (lysin), antigen, complement, and sensitized sheep rbc are required.

-Complement is fixed to a Ab-Ag-complex.

-Fixed complement cannot participate in rbc lysis positive reaction or identification .

Complement fixation test

Nephelometry ( Turbidimetry )

- Nephelometry is a technique used in immunology to determine levels of IgM, IgG, and IgA.

Immunofluorescence

-A monoclonal antibody labeled with a fluorescent dye:

-Direct testing.

-Indirect testing.

-An example of direct and indirect immunofluorescence testing.

Immunofluorescence testing

Common assays

-Radioimmunoassay (RIA).

-Enzyme-Linked Immunosorbent Assay (ELISA).

Radioimmunoassay (RIA)

-Antibody or antigen is labeled.

-Detect corresponding antibody or antigen.

-Competition between unlabeled and labeled molecule.

Radioimmunoassay (RIA) : ( Continued )

Enzyme-Linked Immunosorbent Assay (ELISA)

-Safer & less costly than RIAs.

-ELISA methods can detect any infectious disease provided if we have antibodies and antigen to any infection, enzyme or any substance.

Enzyme-Linked Immunosorbent Assay (ELISA)

-Horseradish peroxidase or alkaline phosphatase.

-Enzyme releases dye (chromogen).

-Types of test:

-Indirect ( Non-competitive ).

-Capture or Sandwich.

-Competitive ELISA.

Ex: detecting the presence of serum AbsAbs against HIV

An example of the indirect ELISA method

An example of the capture ELISA method

An example on competitive ELISA method