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    Some Immunological Test: Presented by Alaa Faeiz Ashwaaq Dyaa Aseel Abd AL-Razaq Supervised by D.Feras

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    Chernet Tuge
    This document summarizes several types of immunological tests used to detect antigens and antibodies, including: 1) Precipitation and agglutination tests detect the formation of aggregates when antigens and antibodies interact. Precipitation tests detect insoluble complexes, while agglutination tests detect interactions between particulate or coated antigens and multivalent antibodies. 2) ELISA tests use enzyme-linked antibodies to detect antigens or antibodies bound to a well. Direct ELISAs detect antigens, while indirect ELISAs detect antibodies in a sample. 3) Other tests mentioned include immunofluorescence assays, radioimmunoassays, latex agglutination tests, haemagglutination tests, and tube agglutination tests like the

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    Some Immunological Test: Presented by Alaa Faeiz Ashwaaq Dyaa Aseel Abd AL-Razaq Supervised by D.Feras

    Uploaded by

    Chernet Tuge
    44 views33 pages
    This document summarizes several types of immunological tests used to detect antigens and antibodies, including: 1) Precipitation and agglutination tests detect the formation of aggregates when antigens and antibodies interact. Precipitation tests detect insoluble complexes, while agglutination tests detect interactions between particulate or coated antigens and multivalent antibodies. 2) ELISA tests use enzyme-linked antibodies to detect antigens or antibodies bound to a well. Direct ELISAs detect antigens, while indirect ELISAs detect antibodies in a sample. 3) Other tests mentioned include immunofluorescence assays, radioimmunoassays, latex agglutination tests, haemagglutination tests, and tube agglutination tests like the

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    This document summarizes several types of immunological tests used to detect antigens and antibodies, including: 1) Precipitation and agglutination tests detect the formation of aggregates when antigens and antibodies interact. Precipitation tests detect insoluble complexes, while agglutination tests detect interactions between particulate or coated antigens and multivalent antibodies. 2) ELISA tests use enzyme-linked antibodies to detect antigens or antibodies bound to a well. Direct ELISAs detect antigens, while indirect ELISAs detect antibodies in a sample. 3) Other tests mentioned include immunofluorescence assays, radioimmunoassays, latex agglutination tests, haemagglutination tests, and tube agglutination tests like the

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    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    44 views33 pages

    Some Immunological Test: Presented by Alaa Faeiz Ashwaaq Dyaa Aseel Abd AL-Razaq Supervised by D.Feras

    Uploaded by

    Chernet Tuge
    This document summarizes several types of immunological tests used to detect antigens and antibodies, including: 1) Precipitation and agglutination tests detect the formation of aggregates when antigens and antibodies interact. Precipitation tests detect insoluble complexes, while agglutination tests detect interactions between particulate or coated antigens and multivalent antibodies. 2) ELISA tests use enzyme-linked antibodies to detect antigens or antibodies bound to a well. Direct ELISAs detect antigens, while indirect ELISAs detect antibodies in a sample. 3) Other tests mentioned include immunofluorescence assays, radioimmunoassays, latex agglutination tests, haemagglutination tests, and tube agglutination tests like the

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    Some Immunological Test

    Presented by
    Alaa Faeiz
    Ashwaaq Dyaa
    Aseel Abd AL-Razaq
    Supervised by D.Feras
    Alaa Faeiz
    Antigen -Antibody Reactions .

    • Antigen – antibody reactions are performed to


    determine the presence of either the antigen or
    antibody. ( serological tests ).

    • One of the two components has to be known.

    • e.g. with a known antigen, such as influenza virus , a


    test can determine whether antibody to the virus is
    present or not .
    There are different types of antigen antibody
    ractions.
    Include:
    - Precipitation reactions(single and double)
    - Agglutination reactions(latex,
    haemagglutination and bacterial Ags )
    -Labeled Antibody serology include:
    -ELISA
    -Immuno-fluorescence assay
    - Radio Immuno Assay (RIA)
    Precipitation test :

    • In this test antigen is in soluble form (solution).

    • Antibody cross -links antigen molecules to form


    aggregates (precipitates) in the zone of
    equivalence:at optimal proportion of antigen and
    antibody.

    • Precipitation test can be performed in solution or in


    semi- solid medium (agar).
    Precipitation in Agar.

    • Single radial immunodiffusion:(MANCINI TEST)

    • Antibody is incorporated into agar and antigen


    introduced into the well.

    • As antigen diffuses into agar precipitation rings form


    depending on the concentration of the antigen.
    • This technique is commonly used for determining the
    concentrations of various plasma proteins such as IgG,
    and IgM in patients suspected to be suffering from
    agamma-globulinaemia and multiple myeloma,
    respectively and others.
    Single Radial Immunodiffusion.
    Double immunodiffusion.(Ouchterlony analysis)

    • Antigen and antibody are placed in different wells


    in agar and allowed to diffuse and form
    precipitation lines at the points of optimal
    concentrations.

    • This method is used to determine whether


    antigens are related, identical or non –
    identical.
    Double immunodiffusion
    Ashwaaq Dyaa
    Agglutination

    • In this test the antigen is particulate (e.g. bacteria and red


    blood cells) or an inert particle (latex beads) coated
    with antigen.

    • Antibody is divalent and cross links the multivalent antigen


    to form a lattice network or clumps
    (agglutination).

    • This reaction can be performed in a tube or on a glass


    slide e.g. ABO blood grouping.
    Latex agglutination
    • This test is a laboratory method to check for
    certain antibodies or antigens in a variety of
    bodily fluids including saliva, urine,
    cerebrospinal fluid, or blood. Examples of
    tests based on latex agglutination reaction
    include pregnancy test, C-reactive protein,,
    and IgM rheumatoid factors and others.
    • the sample is mixed
    with latex beads
    coated with a
    specific antibody or
    antigen. If the
    suspected substance
    is present, the latex
    beads will clump
    together
    (agglutinate)
    Antigen Antibody Reactions

    • Haemaggultination
    Tests:
    The clumping or clustering
    of red blood cells caused by
    certain viruses, antibodies, or
    other substances.
    A.Simple haemagglutination
    : when the red blood cells
    agglutinated by anti sera
    directed to them.
    • B.passive
    haemagglutination
    Red cells can also
    absorb many
    antigens and when
    mixed with specific
    antibodies will form
    clumps i.e. red cells
    are passive carriers .
    Agglutination test of suspensions
    The most common example of this category
    is “WIDAL TEST”
    Widal test is a tube agglutination test
    employed in the serological diagnosis of
    enteric fever . Patients' suffering from
    enteric fever would posses antibodies in
    their sera which can react and agglutinate
    serial doubling dilutions of killed , coloured
    salmonella antigens in a tube agglutination
    test .
    A. Rapid screening test
    1)Place 2 drops of patient's serum .
    2)Add 1 drop of the suspension (S.typhi and
    S.paratyphi ,A and B for H and O antigen ) .
    3)Mix and read within 1 minute .
    • B. tube test, also known as a culture tube or sample
    tube, is a common piece of laboratory glassware
    consisting of a finger-like length of glass or clear plastic
    tubing, open at the top, usually with a rounded U-shaped
    bottom. Tube agglutination test fordetermining antibody
    titer
    The procedure is done as follows :
    1)Dilute patient serum in normal saline
    (1\20 ,1\40 ,1\80 1\160 ,1\320 ,1\640
    ,1\1280).
    2)Add 1 drop of the suspension.
    3)Mix and incubate (O antigen at 50 °C
    for 4hrs ,while H antigen at 50 °C for
    2hrs) .
    4)Examine for agglutination at the bottom
    of the tube.
    c.Rapid slide titration
    Similar to rapid screening test ,but known
    quantities of serum are added (e.g. 80 ul, 40
    ul, 20 ul, 10 ul, 5 ul, 1.0 give dilution of 1\20 ,
    1\40 , 1\80 , 1\160 , 1\320 respectively) .
    Aseel Abd AL-Razaq
    Enzyme Linked Immunosorbent Assay (ELISA)

    • has become one of the most widely used serological tests for
    antibody or antigen detection.
    • ELISA include:
    • Direct and Indirect immunosorbent assay .
    • The direct ELISA is used to detect antigens against a specific
    antibody bound in a test well.
    • The indirect ELISA is used to detect antibodies against an antigen
    bound in a test well.
    • ELISA techniques use antibodies linked to an enzyme and
    Antigen- antibody reactions are detected by enzyme activity.
    Direct ELISA
    • Direct ELISA for antigen detection.
    • A common use of the direct ELISA test is to detect the
    presence of drugs in urine. For this test,antibodies specific
    for the drug are adsorbed to the well on the microtiter
    plate. When the patient's urine sample is added
    to the well, any of the drug that it contained would bind to
    the antibody and is captured . The well is rinsed to remove
    any unbound drug.
    • To make a visible test, more antibodies specific to the drug
    are now added (these antibodies have an enzyme attached
    to them- therefore, the term enzyme-linked ) and will react
    with the already-captured drug, forming a "sandwich" of
    antibody/drug/enzyme-linked antibody This positive test
    can be detected by adding a substrate for the linked
    enzyme; a visible color is produced by the enzyme reacting
    with its substrate and then we measure by
    spectrophotometry.
    Direct ELISA
    ELISA
    Intensity of color correspond to
    concentration of antibody.
    Indirect ELISA

    • The indirect ELISA test, detects antibodies


    in a patient's sample rather than an antigen
    • Indirect ELI SA tests are used, for example,
    to screen blood for antibodies to HIV .For
    such a purpose, the microtiter well contains
    an antigen, such as the inactivated virus that
    causes the disease the test is designed to
    diagnose,
    • A sample of the patient's blood is added to
    the well; if it contains antibodies against the
    virus, they will react with the it. The well is
    rinsed to remove unbound antibodies, If
    antibodies in the blood and the virus in the
    well have attached to each other, they will
    remain in the well-a positive test,
    • To make a positive test visible, some anti-H ISG (an
    immunoglobulin that will attach to any antibody, including
    the one in the patient's serum that has attached to the virus
    in the well)is added, The anti-HISG is linked to an
    enzyme, A positive test consists of a "sandwich" or a
    virus/antibody/enzyme-linked-anti-HISG, At this point, the
    substrate for the enzyme is added, and a positive test is
    detected by the color change caused by the enzyme linked
    to the anti-HISG.
    Indirect ELISA
    Thank You

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